_:b419252316 . _:b6461393 "First, fewer cells expressed lacZ in the CD4\u2212CD8\u2212CD44+CD25\u2212 DN1 compartment of Gata3z/\u2212 (null mutant) ES chimeras than in Gata3z/+ (heterozygous mutant) knockin mice (Hendriks et al., >>1999<<). However, because the DN1 population is heterogeneous and only a very small fraction therein has T lineage potential, in retrospect this experiment was interesting although the final analysis was inconclusive. Second, the generation of" . . . . . _:b6461343 . _:b419252181 . _:b419252170 . . "10.1084%2Fjem.20090934" . . _:b419252275 . . . . _:b419252283 . . . _:b419252212 . _:b6461428 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., >>2008<<), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007) are all known to be important for the generation of ETPs and/or their progenitors, much more extensive characterization will be required to validate the" . . . _:b6461346 . . _:b6461389 . . _:b419252233 . . . _:b419252196 . . _:b6461401 . _:b419252260 . . _:b419252397 . _:b6461416 "When Lin\u2212c-Kit+ fetal liver cells are cultured on OP9-DL1 cells, Notch target genes are induced on day 1, whereas in contrast, GATA-3 was induced only at day 3 in synchrony with CD25, pT\u03B1, and CD3 (Taghon et al., >>2005<<). Although examples exist showing that Notch signaling controls GATA-3 expression at other stages, for example in Th2 cells (Amsen et al., 2007; Fang et al., 2007), in contrast, expression of GATA-3 in bone marrow LSK progenitors actually" . . . . _:b419252242 . . . _:b419252177 . _:b6461385 "as Flt3hiLSK cells) in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., 2005; Lai and Kondo, 2007; Schwarz et al., >>2007<<). Because peripheral myeloid and B lymphoid cells develop normally in Gata3z/z hematopoietic chimeras (Fig." . _:b419252376 . _:b419252222 . _:b419252372 . . _:b419252263 . _:b6461358 "Because B lymphoid cells develop normally in OP9 co-culture of Gata3 null fetal liver cells (Hozumi et al., >>2008<<), it was anticipated that other hematopoietic lineages would develop normally in Gata3g/g embryos." . . . _:b6461385 . _:b6461363 "Although MPPs have been characterized as the most efficient T cell progenitors at the adult stage (Schwarz and Bhandoola, 2004; Schwarz et al., >>2007<<), the precise characteristics and phenotype of the most efficient prethymic T progenitor cells during fetal development are unknown." . _:b419252190 . . . . _:b419252244 . _:b419252253 . . . _:b6461423 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., >>2007<<; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007)" . . . _:b6461341 "Progenitors in the bone marrow were analyzed as previously described (Sambandam et al., >>2005<<; Rumfelt et al., 2006)." . _:b419252382 . _:b419252371 . . _:b6461342 . _:b419252278 . . _:b419252165 . _:b419252183 . . . . . _:b419252198 . . _:b419252247 . _:b419252398 . _:b6461401 "LMPPs in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., 2005; Lai and Kondo, 2007; Schwarz et al., >>2007<<). One immediate possibility is that GATA-3 is required for the homing step from the fetal liver or bone marrow to the thymus. We feel that this is unlikely to be a primary cause of the reduction in the number of ETPs and later stage T" . _:b419252374 . . . _:b419252390 . _:b6461381 . _:b6461361 "The first wave of T progenitor cell migration out of the embryonic fetal liver into the thymic primordium is observed at around e12 to e13 in mice (Owen and Ritter, 1969; Jotereau et al., >>1987<<). Although MPPs have been characterized as the most efficient T cell progenitors at the adult stage (Schwarz and Bhandoola, 2004; Schwarz et al., 2007), the precise characteristics and phenotype of the most efficient prethymic T" . _:b419252304 . . . _:b419252319 . _:b419252238 . _:b419252343 . _:b419252252 . _:b6461353 . . . . . _:b6461389 "To independently confirm the requirement for GATA-3 in the development of adult ETPs, we intercrossed Gata3 conditional knockout (Gata3flox) mice (Kurek et al., >>2007<<) with mice bearing the Mx1cre transgene (K\u00FChn et al., 1995), followed by induction of Cre recombinase by pI-pC." . . . . _:b6461345 "It should be noted that the B220 epitope is expressed on early to mature B lineage cells as well as on some activated T, dendritic, and NK cells (Watanabe and Akaike, >>1994<<; Huntington et al., 2007)." . . _:b6461398 "LMPPs in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, >>2004<<; Adolfsson et al., 2005; Lai and Kondo, 2007; Schwarz et al., 2007)." . . _:b419252250 . _:b6461395 "Finally, the progeny of Gata3 null mutant progenitors was significantly reduced in fetal thymus organ culture (Hozumi et al., >>2008<<), underscoring the likelihood of a cell-autonomous role for GATA-3 function in T lineage development." . _:b6461360 . . _:b6461429 . _:b6461380 . . _:b419252340 . _:b6461384 . _:b6461407 . . _:b419252200 . . . _:b419252158 . . _:b6461350 . . _:b419252348 . . _:b419252414 . _:b419252352 . _:b419252358 . _:b419252338 . . _:b6461366 "T lineage\u2013restricted progenitors in the fetal liver have been reported (Rodewald et al., 1994; Kawamoto et al., 1997) and have been described within the Lin\u2212c-KithiIL-7R\u03B1+paired immunoglobulin receptor (PIR)+ fraction (Masuda et al., >>2005<<). Of particular note, the developmental relationship between fetal prethymic T-restricted progenitors and fetal thymic ETPs is presently obscure." . _:b419252172 . . _:b6461388 . . _:b6461356 . . . . _:b419252300 . _:b6461400 . _:b419252158 "11"^^ . _:b419252159 "9"^^ . _:b6461357 "Linlo population in Gata3g/g mice, because gating on Linlo thymocytes had been shown to enrich for bona fide ETPs that are found in the complex hematopoietic compartment described by the CD44+CD25\u2212 DN1 population (Sambandam et al., >>2005<<). As shown in Fig. S3 (A\u2013C), the absolute number of LinloCD44+CD25\u2212 cells per thymus was also reduced (to 21.9% [e15.5] or 14% [e18.5] of wild-type controls)." . _:b6461402 . _:b6461427 . _:b419252396 . _:b419252166 "6"^^ . _:b6461369 . _:b419252167 "6"^^ . _:b419252409 . _:b6461341 . _:b6461400 "LMPPs in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., 2005; Lai and Kondo, >>2007<<; Schwarz et al., 2007)." . _:b6461405 "In the absence of GATA-3, bone marrow immature NK cells develop normally, whereas thymic NK cells are not generated (Samson et al., 2003; Vosshenrich et al., >>2006<<). These data suggest a function for GATA-3 in a lymphocyte progenitor population after thymic entry and exposure to Notch ligands. A second possibility is that GATA-3 activity is required at the very beginning of ETP generation, a" . _:b419252164 "7"^^ . . _:b419252165 "6"^^ . _:b419252346 . _:b419252162 "7"^^ . _:b419252163 "7"^^ . _:b419252240 . _:b419252160 "7"^^ . . _:b419252185 . _:b419252342 . _:b419252161 "7"^^ . . . _:b419252173 "5"^^ . _:b419252202 . . _:b419252245 . . _:b419252172 "5"^^ . _:b419252267 . _:b419252175 "4"^^ . _:b419252174 "4"^^ . _:b419252170 "6"^^ . _:b419252276 . _:b6461420 . _:b419252168 "6"^^ . _:b6461358 . _:b419252277 . . _:b419252378 . _:b419252171 "5"^^ . _:b419252169 "6"^^ . _:b419252181 "4"^^ . . _:b419252180 "4"^^ . _:b6461357 . _:b419252183 "4"^^ . _:b6461348 . _:b419252182 "4"^^ . . . _:b419252257 . _:b419252177 "4"^^ . . _:b6461427 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., >>2008<<), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007) are all known to be important for the generation of ETPs and/or their progenitors, much more extensive characterization will be" . _:b419252176 "4"^^ . _:b419252158 . _:b419252159 . _:b419252179 "4"^^ . _:b419252412 . _:b6461349 . . _:b419252178 "4"^^ . _:b419252395 . _:b419252189 "3"^^ . _:b6461403 . _:b419252188 "3"^^ . _:b419252354 . _:b419252191 "3"^^ . . _:b419252410 . _:b419252190 "3"^^ . _:b419252294 . _:b419252360 . _:b419252411 . . _:b419252185 "4"^^ . _:b419252391 . _:b419252184 "4"^^ . . . _:b6461424 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., >>2007<<), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007) are all known to be" . _:b419252192 . _:b6461335 . _:b419252187 "4"^^ . _:b419252193 . _:b419252194 . _:b419252195 . _:b419252196 . _:b419252186 "4"^^ . _:b419252197 . _:b419252198 . _:b419252199 . _:b419252197 "3"^^ . _:b419252200 . _:b419252201 . _:b419252202 . _:b6461395 . _:b419252203 . _:b419252196 "3"^^ . _:b419252204 . . _:b419252205 . _:b419252206 . _:b419252207 . _:b419252199 "3"^^ . _:b419252208 . . _:b419252209 . _:b419252210 . _:b419252211 . _:b419252198 "3"^^ . _:b419252212 . _:b419252213 . _:b419252214 . . _:b419252215 . _:b419252193 "3"^^ . _:b419252216 . . _:b6461394 . _:b419252217 . _:b419252218 . _:b419252219 . _:b419252192 "3"^^ . _:b419252220 . _:b419252221 . _:b419252222 . _:b419252223 . _:b6461410 . _:b419252195 "3"^^ . _:b419252160 . _:b419252161 . . _:b419252162 . _:b419252163 . _:b419252194 "3"^^ . _:b419252164 . _:b419252165 . _:b419252166 . _:b419252167 . _:b419252205 "3"^^ . _:b419252168 . _:b419252169 . _:b419252170 . _:b419252171 . _:b419252204 "3"^^ . _:b419252172 . _:b419252173 . _:b419252204 . _:b419252174 . _:b419252175 . _:b419252207 "3"^^ . _:b419252176 . . _:b419252177 . _:b419252178 . _:b419252179 . _:b6461418 "Although examples exist showing that Notch signaling controls GATA-3 expression at other stages, for example in Th2 cells (Amsen et al., 2007; Fang et al., >>2007<<), in contrast, expression of GATA-3 in bone marrow LSK progenitors actually precedes the activation of Notch signaling (Sambandam et al., 2005; Lai and Kondo, 2007), and Notch signaling is dispensable for HSC function in adult mice" . _:b419252206 "3"^^ . . _:b419252180 . _:b419252181 . _:b419252182 . _:b419252183 . _:b419252201 "3"^^ . _:b419252184 . _:b419252185 . _:b419252186 . _:b419252187 . _:b419252200 "3"^^ . _:b419252188 . _:b419252189 . _:b419252190 . _:b419252375 . _:b419252191 . _:b419252203 "3"^^ . _:b419252202 "3"^^ . . _:b6461379 . . . _:b419252213 "3"^^ . _:b6461344 . . _:b419252212 "3"^^ . _:b6461410 "1884), and GATA-3 activity in the first through the fourth arches is required for the development of several different organs derived from the pharyngeal arch mesenchyme during early embryogenesis (unpublished data; Lim et al., >>2000<<)." . _:b419252234 . _:b419252215 "3"^^ . _:b419252214 "3"^^ . _:b419252166 . _:b6461425 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., >>2007<<), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007) are all known to be important for the generation of ETPs and/or" . _:b419252209 "3"^^ . _:b419252208 "3"^^ . _:b419252211 "3"^^ . _:b419252297 . . _:b419252210 "3"^^ . _:b6461350 "To examine the developmental potential of Gata3 null mutant (lacZ knockin, Gata3z/z; van Doorninck et al., >>1999<<) T cell progenitors in vitro, we isolated and co-cultured fetal liver cells from e12.5/e13.5 catecholamine-rescued Gata3z/z embryos on OP9-DL1 cells (Schmitt and Z\u00FA\u00F1iga-Pfl\u00FCcker, 2002) to promote T cell development." . _:b419252394 . _:b419252221 "3"^^ . _:b419252210 . _:b6461390 "To independently confirm the requirement for GATA-3 in the development of adult ETPs, we intercrossed Gata3 conditional knockout (Gata3flox) mice (Kurek et al., 2007) with mice bearing the Mx1cre transgene (K\u00FChn et al., >>1995<<), followed by induction of Cre recombinase by pI-pC. Thymocytes from compound mutant Gata3flox/flox:" . _:b419252220 "3"^^ . . . _:b419252223 "3"^^ . . _:b419252222 "3"^^ . _:b419252217 "3"^^ . _:b419252216 "3"^^ . _:b419252219 "3"^^ . . _:b6461364 . _:b419252229 "3"^^ . _:b419252218 "3"^^ . _:b419252228 "3"^^ . _:b419252231 "3"^^ . . . _:b419252230 "3"^^ . . _:b419252225 "3"^^ . _:b419252323 . _:b419252345 . _:b6461430 "Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., >>2007<<) are all known to be important for the generation of ETPs and/or their progenitors, much more extensive characterization will be required to validate the relationships between and epistatic hierarchy among these factors during the" . _:b419252224 "3"^^ . _:b419252227 "3"^^ . _:b419252321 . _:b419252226 "3"^^ . _:b419252237 "3"^^ . _:b419252236 "3"^^ . _:b419252317 . _:b419252239 "3"^^ . . _:b419252322 . _:b419252238 "3"^^ . _:b419252233 "3"^^ . . . _:b419252232 "3"^^ . _:b419252236 . _:b419252235 "3"^^ . _:b6461414 . _:b419252271 . _:b419252363 . _:b419252234 "3"^^ . _:b6461409 . _:b419252245 "2"^^ . _:b419252209 . _:b419252244 "2"^^ . _:b419252305 . _:b6461411 . _:b6461348 "Although diminished GATA-3 activity in fetal liver cells was reported to result in a loss of Thy-1+ T cells after expansion in fetal thymus organ culture (Hattori et al., >>1996<<; Hozumi et al., 2008), the specific affected stages remained unclear." . _:b419252247 "2"^^ . . _:b419252246 "2"^^ . _:b419252361 . . _:b419252241 "3"^^ . . _:b419252240 "3"^^ . _:b419252243 "3"^^ . _:b6461422 . _:b6461407 "Because Flt3-expressing early ETPs retain B, myeloid, dendritic, and NK cell potential (Porritt et al., >>2004<<; Balciunaite et al., 2005; Sambandam et al., 2005), it is possible that TSP preferentially develops to these non\u2013T lineages in Gata3 mutants." . _:b419252298 . _:b6461415 . _:b419252242 "3"^^ . . _:b419252253 "2"^^ . . . _:b6461426 . . _:b419252252 "2"^^ . _:b6461419 . _:b419252255 "2"^^ . _:b6461368 "Although it is known that the adult and fetal stages of T cell development share similarities, they nonetheless differ in numerous and well-defined features (for reviews see Kincade et al., 2002; David-Fung et al., >>2006<<). We previously used a Gata3 lacZ knockin allele that has proven to be an extremely useful tool to monitor cells that express GATA-3 (van Doorninck et al., 1999; Lim et al., 2000)." . _:b419252254 "2"^^ . . _:b419252249 "2"^^ . . _:b419252248 "2"^^ . . _:b419252251 "2"^^ . _:b419252334 . _:b419252250 "2"^^ . _:b419252417 . _:b419252207 . _:b419252261 "2"^^ . _:b419252260 "2"^^ . . _:b419252263 "2"^^ . . . . . _:b419252262 "2"^^ . . _:b419252257 "2"^^ . . _:b6461382 . _:b419252256 "2"^^ . . _:b419252225 . _:b6461426 "Although the Flt3 cytokine receptor (Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, >>2004<<; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity (Talebian et al., 2007) are all known to be important for the generation of ETPs and/or their progenitors, much more extensive" . _:b419252259 "2"^^ . . _:b419252265 . _:b419252258 "2"^^ . _:b419252269 "2"^^ . _:b419252268 "2"^^ . . _:b419252271 "2"^^ . _:b419252270 "2"^^ . . _:b6461337 . _:b419252265 "2"^^ . . . _:b419252264 "2"^^ . _:b419252267 "2"^^ . _:b6461352 . _:b6461338 "Grosveld (Erasmus Medical Center, Rotterdam, Netherlands; Kurek et al., >>2007<<). C57BL/6-Ly5B6 (CD45.2) mice and C57BL/6-Ly5SJL (CD45.1) mice were purchased from the Jackson Laboratory." . _:b419252227 . . _:b419252266 "2"^^ . _:b419252351 . _:b419252277 "2"^^ . . . _:b6461404 "In the absence of GATA-3, bone marrow immature NK cells develop normally, whereas thymic NK cells are not generated (Samson et al., >>2003<<; Vosshenrich et al., 2006)." . _:b419252276 "2"^^ . _:b419252279 "2"^^ . _:b6461352 "Gata3 null mutant mice fail to generate even a rudimentary thymus (Lim et al., >>2000<<; Moriguchi et al., 2006), whereas Gata3 conditionally mutant mice that were crossed to mice bearing an lck-directed Cre transgene, and therefore lacked Gata3 at the DN3 stage and thereafter, were reported to have normal numbers of" . _:b419252278 "2"^^ . _:b419252273 "2"^^ . _:b419252355 . _:b6461398 . _:b419252272 "2"^^ . _:b419252275 "2"^^ . _:b419252274 "2"^^ . _:b419252344 . "PMC0" . . _:b419252285 "2"^^ . _:b419252159 . _:b419252243 . _:b419252284 "2"^^ . _:b419252180 . _:b419252287 "2"^^ . _:b419252366 . _:b419252286 "2"^^ . _:b419252281 "2"^^ . . . _:b419252280 "2"^^ . _:b419252241 . . _:b419252283 "2"^^ . _:b6461334 "materials and methods" . _:b419252282 "2"^^ . . _:b419252293 "2"^^ . _:b6461370 . _:b6461355 . _:b419252318 . _:b419252292 "2"^^ . _:b419252295 "2"^^ . _:b6461356 "Forced expression of GATA-3 was previously shown to induce c-Kit expression (Anderson et al., 2002; Taghon et al., >>2007<<). To ask whether the observed reduction in the number of ETPs and DN2 cells might be caused by reduced c-Kit expression, we analyzed CD44 and CD25 within the Linlo population in Gata3g/g mice, because gating on Linlo thymocytes had been" . _:b419252273 . . _:b419252294 "2"^^ . . . _:b419252289 "2"^^ . . _:b419252288 "2"^^ . _:b419252315 . _:b419252291 "2"^^ . . . . . . _:b419252290 "2"^^ . _:b419252266 . _:b419252301 "2"^^ . _:b419252418 . . _:b419252413 . _:b419252353 . _:b419252300 "2"^^ . . _:b419252303 "2"^^ . _:b419252302 "2"^^ . _:b419252297 "2"^^ . _:b419252296 "2"^^ . _:b419252383 . _:b419252299 "2"^^ . . _:b419252298 "2"^^ . _:b419252384 . . _:b419252309 "2"^^ . _:b419252308 "2"^^ . . . _:b419252341 . _:b419252311 "2"^^ . _:b6461382 "Lymphoid-primed MPPs (LMPPs; defined as Flt3hiLSK cells) in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, >>2004<<; Adolfsson et al., 2005; Lai and Kondo, 2007; Schwarz et al., 2007). Because peripheral myeloid and B lymphoid cells develop normally in Gata3z/z hematopoietic chimeras (Fig." . _:b419252310 "2"^^ . _:b419252305 "2"^^ . _:b419252304 "2"^^ . _:b419252307 "2"^^ . . . _:b419252306 "2"^^ . _:b419252317 "2"^^ . . . _:b6461362 . _:b419252316 "2"^^ . _:b419252281 . _:b419252319 "2"^^ . _:b419252318 "2"^^ . _:b419252296 . _:b419252313 "2"^^ . . _:b419252312 "2"^^ . _:b419252315 "2"^^ . _:b419252239 . . _:b419252288 . _:b419252314 "2"^^ . . _:b419252325 "2"^^ . _:b6461392 _:b6461412 . _:b6461392 _:b6461413 . _:b6461392 _:b6461414 . _:b6461392 _:b6461415 . _:b419252324 "2"^^ . _:b6461392 _:b6461408 . _:b6461392 _:b6461409 . _:b419252224 . _:b6461392 _:b6461410 . _:b6461392 _:b6461411 . _:b419252327 "2"^^ . _:b6461392 _:b6461420 . _:b6461392 _:b6461421 . _:b419252289 . _:b6461392 _:b6461422 . _:b6461392 _:b6461423 . _:b419252326 "2"^^ . _:b6461392 _:b6461416 . _:b6461392 _:b6461417 . _:b6461392 _:b6461418 . _:b6461392 _:b6461419 . _:b419252321 "2"^^ . _:b6461392 _:b6461428 . . _:b6461392 _:b6461429 . _:b6461392 _:b6461430 . _:b6461367 "Although it is known that the adult and fetal stages of T cell development share similarities, they nonetheless differ in numerous and well-defined features (for reviews see Kincade et al., >>2002<<; David-Fung et al., 2006). We previously used a Gata3 lacZ knockin allele that has proven to be an extremely useful tool to monitor cells that express GATA-3 (van Doorninck et al., 1999; Lim et al., 2000)." . . _:b419252320 "2"^^ . _:b6461392 _:b6461424 . _:b6461392 _:b6461425 . _:b419252270 . _:b6461392 _:b6461426 . _:b419252287 . . . _:b6461392 _:b6461427 . _:b419252323 "2"^^ . _:b419252369 . _:b419252322 "2"^^ . _:b419252347 . . _:b419252333 "2"^^ . . _:b419252332 "2"^^ . _:b419252335 "2"^^ . _:b6461428 . . _:b419252334 "2"^^ . . _:b419252329 "2"^^ . _:b6461392 _:b6461396 . _:b6461392 _:b6461397 . _:b6461392 _:b6461398 . _:b6461392 _:b6461399 . _:b419252328 "2"^^ . _:b6461392 _:b6461393 . _:b6461392 _:b6461394 . _:b6461392 _:b6461395 . _:b419252331 "2"^^ . _:b419252211 . _:b6461392 _:b6461404 . _:b6461392 _:b6461405 . _:b6461392 _:b6461406 . _:b6461392 _:b6461407 . _:b419252330 "2"^^ . _:b6461392 _:b6461400 . _:b6461392 _:b6461401 . _:b6461392 _:b6461402 . _:b6461392 _:b6461403 . _:b419252341 "2"^^ . . _:b419252256 . . . _:b419252257 . _:b419252258 . _:b419252340 "2"^^ . _:b419252259 . . _:b419252260 . _:b419252261 . _:b419252262 . _:b419252343 "2"^^ . _:b419252263 . _:b419252264 . _:b419252265 . _:b419252266 . _:b419252342 "2"^^ . _:b419252267 . _:b419252268 . _:b419252269 . _:b419252217 . _:b419252270 . _:b419252337 "2"^^ . _:b419252271 . _:b419252272 . _:b419252273 . . _:b419252274 . _:b419252336 "2"^^ . _:b419252275 . _:b419252276 . _:b419252230 . _:b419252277 . _:b419252278 . _:b419252339 "2"^^ . _:b419252279 . _:b419252280 . _:b419252281 . _:b419252282 . _:b419252338 "2"^^ . _:b419252283 . _:b419252284 . _:b419252285 . _:b419252286 . _:b419252359 . _:b419252287 . _:b419252349 "2"^^ . _:b419252224 . _:b419252225 . _:b419252226 . . _:b419252227 . _:b419252348 "2"^^ . _:b419252228 . _:b419252229 . _:b6461413 . _:b419252230 . _:b419252351 "2"^^ . _:b6461419 "expression at other stages, for example in Th2 cells (Amsen et al., 2007; Fang et al., 2007), in contrast, expression of GATA-3 in bone marrow LSK progenitors actually precedes the activation of Notch signaling (Sambandam et al., >>2005<<; Lai and Kondo, 2007), and Notch signaling is dispensable for HSC function in adult mice (Maillard et al., 2008)." . _:b419252231 . _:b419252232 . _:b6461412 . . _:b419252233 . _:b419252234 . _:b419252350 "2"^^ . _:b419252235 . _:b419252236 . _:b419252389 . _:b419252237 . _:b419252238 . _:b6461355 "Forced expression of GATA-3 was previously shown to induce c-Kit expression (Anderson et al., >>2002<<; Taghon et al., 2007)." . _:b419252345 "2"^^ . _:b419252239 . . _:b6461383 "Lymphoid-primed MPPs (LMPPs; defined as Flt3hiLSK cells) in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., >>2005<<; Lai and Kondo, 2007; Schwarz et al., 2007). Because peripheral myeloid and B lymphoid cells develop normally in Gata3z/z hematopoietic chimeras (Fig." . _:b419252240 . _:b419252241 . _:b419252242 . _:b419252280 . _:b419252243 . _:b419252344 "2"^^ . _:b419252244 . _:b419252245 . _:b419252246 . _:b419252347 "2"^^ . _:b419252247 . . _:b419252248 . _:b419252249 . . _:b419252250 . _:b419252346 "2"^^ . _:b419252251 . _:b419252252 . _:b419252253 . _:b419252254 . _:b419252357 "2"^^ . _:b419252255 . _:b419252320 . _:b419252321 . _:b419252322 . _:b419252356 "2"^^ . _:b419252323 . . _:b419252324 . _:b419252325 . _:b419252326 . _:b419252359 "2"^^ . _:b419252327 . _:b419252328 . _:b419252329 . _:b6461421 . _:b419252330 . _:b419252358 "2"^^ . _:b419252331 . _:b419252332 . _:b419252333 . _:b419252334 . . _:b419252335 . _:b419252353 "2"^^ . _:b419252336 . _:b419252337 . _:b419252338 . _:b419252352 "2"^^ . _:b419252339 . _:b419252340 . _:b419252341 . _:b419252342 . _:b419252355 "2"^^ . _:b419252343 . _:b419252344 . _:b419252345 . . _:b419252346 . _:b419252354 "2"^^ . _:b419252347 . _:b419252348 . . _:b419252349 . _:b419252350 . _:b419252365 "2"^^ . _:b419252351 . _:b419252288 . _:b419252289 . _:b419252290 . _:b419252364 "2"^^ . _:b419252291 . _:b419252292 . _:b6461397 "paper, we show that the vast majority of GATA-3\u2013expressing cells in the DN1 compartment also expressed NK1.1, which is abundantly expressed on c-Kit\u2212 cells of the CD4\u2212CD8\u2212CD44+ population (Laurent et al., 2004; Balciunaite et al., >>2005<<). This observation suggested that a detailed reevaluation of the contributions of GATA-3 to early T cell development was warranted." . _:b6461377 "5 A, GFP fluorescence in the DN, DP, CD4 SP, and CD8 SP thymocytes isolated from 6-wk-old Gata3g/+ mice was generally reflective of the reported profile for wild-type GATA-3 expression (Hendriks et al., >>1999<<). As anticipated, a majority of the NK1.1+ and \u03B3\u03B4TCR+ cells expressed abundant GATA-3.eGFP fusion protein. Detailed analysis of the complex DN1 population revealed that (on average) 83.1 \u00B1 2.9% of the eGFP+ cells were also NK1.1+, thus" . _:b419252293 . _:b419252294 . _:b6461369 "We previously used a Gata3 lacZ knockin allele that has proven to be an extremely useful tool to monitor cells that express GATA-3 (van Doorninck et al., >>1999<<; Lim et al., 2000)." . _:b419252367 "2"^^ . _:b419252295 . _:b419252296 . _:b6461371 "We previously used a Gata3 lacZ knockin allele that has proven to be an extremely useful tool to monitor cells that express GATA-3 (van Doorninck et al., 1999; Lim et al., 2000). Hendriks et al. (>>1999<<) found that fewer cells express \u03B2-galactosidase in the classically defined DN1 (CD4\u2212CD8\u2212CD44+CD25\u2212) compartment of Gata3z/\u2212 (null) ES chimeras in comparison to heterozygous Gata3z/+ ES chimeric mice, which originally suggested an early" . _:b419252297 . _:b419252298 . _:b419252366 "2"^^ . _:b419252299 . _:b419252300 . _:b419252301 . . _:b419252302 . _:b419252361 "2"^^ . _:b419252303 . . _:b419252304 . . _:b419252305 . _:b419252306 . _:b419252360 "2"^^ . _:b419252307 . . _:b419252308 . _:b419252309 . _:b419252268 . _:b419252310 . _:b419252213 . _:b419252363 "2"^^ . _:b419252311 . _:b419252312 . _:b419252197 . _:b419252313 . _:b419252314 . _:b419252362 "2"^^ . _:b419252315 . _:b419252316 . _:b419252182 . _:b419252317 . _:b419252318 . _:b419252373 "2"^^ . _:b419252319 . _:b419252384 . _:b419252385 . _:b419252203 . _:b419252386 . _:b419252372 "2"^^ . _:b419252387 . _:b419252388 . _:b419252389 . _:b419252390 . _:b419252375 "2"^^ . _:b419252391 . _:b419252310 . _:b419252392 . _:b419252393 . _:b419252226 . _:b419252394 . _:b419252374 "2"^^ . _:b419252395 . _:b419252396 . _:b419252397 . _:b419252398 . _:b419252369 "2"^^ . _:b419252399 . _:b419252400 . _:b419252401 . _:b419252402 . _:b419252368 "2"^^ . _:b419252403 . _:b419252404 . _:b419252405 . _:b419252335 . _:b419252406 . _:b419252371 "2"^^ . _:b419252407 . . _:b419252408 . _:b419252409 . _:b419252410 . _:b419252370 "2"^^ . _:b419252411 . _:b6461334 _:b6461344 . _:b419252285 . _:b419252220 . _:b6461335 "The Gata3z allele (van Doorninck et al., >>1999<<) and Mx1cre transgenic mice (provided by M. Yamamoto, Tohoku University Medical School, Sendai, Japan; K\u00FChn et al., 1995) have been previously described." . _:b419252412 . . _:b6461334 _:b6461345 . _:b6461334 _:b6461346 . _:b419252413 . _:b419252414 . _:b419252381 "2"^^ . _:b419252415 . _:b6461334 _:b6461340 . _:b6461334 _:b6461341 . _:b419252352 . _:b6461334 _:b6461342 . _:b419252353 . _:b6461334 _:b6461343 . _:b419252354 . _:b419252282 . _:b419252380 "2"^^ . _:b419252355 . _:b6461334 _:b6461336 . _:b6461334 _:b6461337 . _:b419252356 . _:b6461334 _:b6461338 . _:b419252357 . _:b6461334 _:b6461339 . _:b419252358 . _:b419252383 "2"^^ . _:b419252359 . _:b419252360 . _:b6461368 . _:b419252361 . _:b6461334 _:b6461335 . _:b419252362 . _:b419252382 "2"^^ . _:b419252363 . _:b419252364 . _:b419252387 . _:b419252365 . _:b419252366 . _:b419252377 "2"^^ . _:b419252367 . _:b419252368 . _:b6461388 "Because GATA-3 is expressed in these fractions (Akashi et al., 2000; Rumfelt et al., >>2006<<), the developmental potential of Gata3z/z cells in these fractions was examined in adoptive transfer experiments." . _:b419252369 . _:b419252370 . _:b419252376 "2"^^ . _:b419252371 . _:b419252372 . _:b419252373 . _:b419252249 . _:b419252374 . _:b419252379 "2"^^ . _:b419252375 . _:b419252221 . _:b419252376 . _:b419252377 . _:b419252378 . _:b419252378 "2"^^ . _:b6461375 . _:b419252379 . . _:b419252380 . _:b419252381 . _:b419252382 . _:b419252389 "2"^^ . _:b419252383 . _:b419252400 . _:b419252279 . _:b419252388 "2"^^ . _:b6461396 . _:b419252391 "2"^^ . _:b419252269 . . _:b419252390 "2"^^ . _:b419252385 "2"^^ . _:b419252332 . _:b419252384 "2"^^ . _:b6461391 "To limit analyses to lymphoid progenitors, we took advantage of the CD27 cell-surface marker, which is expressed in early lymphoid progenitors through mature T cells (Taghon et al., >>2005<<). The number of total and Lin\u2212 fetal liver cells was identical in the Gata3z/z and Gata3+/+ e14.5 embryos (unpublished data). As shown in Fig. 8 (A and B) and Table I, e14.5 Gata3z/z Lin\u2212 fetal livers developed a reduced number of" . _:b419252387 "2"^^ . _:b419252386 "2"^^ . _:b419252189 . _:b419252397 "2"^^ . _:b419252416 . _:b419252417 . _:b419252418 . _:b419252396 "2"^^ . . . _:b419252399 "2"^^ . . . _:b6461381 "Because GATA-3 null ES cells can contribute to myeloid and B lymphoid lineage cells in blastocyst chimeras (Ting et al., >>1996<<), reconstitution efficiency was periodically monitored in peripheral blood myeloid (Gr1+Mac1+) and B lymphoid (B220+) cells, and was found to be the same for fetal liver donors of both genotypes (Fig." . _:b419252398 "2"^^ . _:b419252191 . _:b419252393 "2"^^ . _:b419252392 "2"^^ . _:b6461387 . . . _:b419252395 "2"^^ . _:b419252327 . _:b419252394 "2"^^ . _:b419252274 . . . _:b419252405 "2"^^ . _:b419252324 . . . _:b419252404 "2"^^ . . . _:b419252407 "2"^^ . _:b419252164 . _:b419252406 "2"^^ . . _:b6461370 "We previously used a Gata3 lacZ knockin allele that has proven to be an extremely useful tool to monitor cells that express GATA-3 (van Doorninck et al., 1999; Lim et al., >>2000<<). Hendriks et al. (1999) found that fewer cells express \u03B2-galactosidase in the classically defined DN1 (CD4\u2212CD8\u2212CD44+CD25\u2212) compartment of Gata3z/\u2212 (null) ES chimeras in comparison to heterozygous Gata3z/+ ES chimeric mice, which" . . _:b419252401 "2"^^ . _:b6461354 . . _:b419252400 "2"^^ . _:b419252403 "2"^^ . _:b419252402 "2"^^ . _:b6461347 _:b6461384 . _:b419252413 "2"^^ . _:b6461347 _:b6461385 . _:b6461347 _:b6461386 . _:b6461347 _:b6461387 . _:b6461347 _:b6461388 . _:b6461359 . _:b419252412 "2"^^ . _:b419252331 . _:b6461347 _:b6461389 . _:b6461347 _:b6461390 . _:b419252284 . _:b6461347 _:b6461391 . _:b419252215 . _:b419252415 "2"^^ . _:b6461347 _:b6461376 . _:b6461347 _:b6461377 . _:b6461347 _:b6461378 . _:b6461347 _:b6461379 . _:b6461347 _:b6461380 . _:b419252414 "2"^^ . _:b6461347 _:b6461381 . _:b6461347 _:b6461382 . _:b6461347 _:b6461383 . _:b419252409 "2"^^ . _:b419252408 "2"^^ . _:b6461409 "Because Flt3-expressing early ETPs retain B, myeloid, dendritic, and NK cell potential (Porritt et al., 2004; Balciunaite et al., 2005; Sambandam et al., >>2005<<), it is possible that TSP preferentially develops to these non\u2013T lineages in Gata3 mutants." . _:b419252231 . _:b419252411 "2"^^ . . _:b6461421 "expression of GATA-3 in bone marrow LSK progenitors actually precedes the activation of Notch signaling (Sambandam et al., 2005; Lai and Kondo, 2007), and Notch signaling is dispensable for HSC function in adult mice (Maillard et al., >>2008<<). Thus, the relationship between GATA-3 and Notch signaling during very early T lymphoid development remains unclear." . _:b419252410 "2"^^ . . _:b6461347 _:b6461352 . _:b6461347 _:b6461353 . . _:b6461347 _:b6461354 . _:b6461347 _:b6461355 . _:b6461347 _:b6461356 . _:b6461347 _:b6461357 . _:b419252255 . _:b6461347 _:b6461358 . _:b6461347 _:b6461359 . _:b6461384 "MPPs (LMPPs; defined as Flt3hiLSK cells) in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., 2005; Lai and Kondo, >>2007<<; Schwarz et al., 2007). Because peripheral myeloid and B lymphoid cells develop normally in Gata3z/z hematopoietic chimeras (Fig." . _:b6461347 _:b6461348 . _:b6461354 "Cre transgene, and therefore lacked Gata3 at the DN3 stage and thereafter, were reported to have normal numbers of DN-stage cells per thymus (DN1 through DN4 total) but reduced numbers of DP, CD4 SP, and CD8 SP cells (Pai et al., >>2003<<). We recently generated a Gata3 hypomorphic mutant (Gata3g/g) mouse that develops a diminutive thymus (unpublished data)." . _:b6461347 _:b6461349 . _:b6461347 _:b6461350 . _:b419252405 . _:b6461347 _:b6461351 . _:b6461347 _:b6461368 . _:b419252417 "2"^^ . _:b6461347 _:b6461369 . _:b6461347 _:b6461370 . _:b6461391 . _:b6461347 _:b6461371 . _:b6461347 _:b6461372 . _:b6461424 . _:b419252416 "2"^^ . _:b419252299 . _:b6461347 _:b6461373 . _:b6461347 _:b6461374 . _:b6461347 _:b6461375 . _:b6461347 _:b6461360 . _:b6461425 . _:b6461347 _:b6461361 . _:b6461347 _:b6461362 . _:b6461347 _:b6461363 . _:b6461347 _:b6461364 . _:b419252418 "2"^^ . _:b6461347 _:b6461365 . _:b6461347 _:b6461366 . _:b6461347 _:b6461367 . . _:b419252259 . _:b419252399 . _:b6461373 "have revealed that DN1 cells are actually quite heterogeneous in nature, and contain a mixture of mature and immature NK, DC, B, and myeloid lineage cells, as well as rare T cell progenitors (Allman et al., 2003; Porritt et al., >>2004<<; Balciunaite et al., 2005; Anderson, 2006). A recent study also revealed that GATA-3 is expressed in and is required for the generation of thymic NK cells (Vosshenrich et al., 2006)." . . _:b419252404 . _:b6461343 "To induce T cell development, 1\u20132 \u00D7 105 total fetal liver cells or Lin-depleted fetal liver cells recovered from one embryo were cultured on a feeder layer of OP9-DL1 cells (Schmitt and Z\u00FA\u00F1iga-Pfl\u00FCcker, >>2002<<) in the presence of 5 ng/ml IL-7 and 5 ng/ml Flt3-ligand (R&D Systems). Partial media change was performed every 3\u20134 d." . _:b419252208 . . _:b419252246 . _:b6461342 "Progenitors in the bone marrow were analyzed as previously described (Sambandam et al., 2005; Rumfelt et al., >>2006<<). A cocktail containing antibodies recognizing TER119, Gr-1, B220, CD19, CD3, and Mac-1 was used to exclude Lin+ cells in the bone marrow for the analysis of HSCs, MPPs, and LMPPs. A cocktail containing antibodies recognizing TER119," . . _:b6461359 "Importantly, these findings reveal a pivotal developmental role in vivo for GATA-3 at a stage of T cell development earlier than DN3, which could not be addressed using the strategy pursued by Pai et al. (>>2003<<)." . _:b419252301 . _:b419252309 . _:b419252216 . _:b419252312 . _:b419252407 . _:b419252258 . . . . _:b419252380 . . _:b419252187 . . _:b6461334 . _:b6461415 "Both GATA-3 and Notch signaling are required for T cell development, whereas either factor alone cannot induce thymocyte development (Hozumi et al., >>2008<<). When Lin\u2212c-Kit+ fetal liver cells are cultured on OP9-DL1 cells, Notch target genes are induced on day 1, whereas in contrast, GATA-3 was induced only at day 3 in synchrony with CD25, pT\u03B1, and CD3 (Taghon et al., 2005). Although" . _:b6461335 . . _:b6461416 . . _:b419252229 . _:b6461422 "Thus, the relationship between GATA-3 and Notch signaling during very early T lymphoid development remains unclear. Although the Flt3 cytokine receptor (Sambandam et al., >>2005<<; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., 2009), and RUNX activity" . . _:b419252228 . _:b6461340 . _:b6461394 "Second, the generation of Thy1+ cells from fetal liver progenitors in fetal thymus organ culture was inhibited by treatment with GATA-3 antisense oligonucleotides (Hattori et al., >>1996<<). Finally, the progeny of Gata3 null mutant progenitors was significantly reduced in fetal thymus organ culture (Hozumi et al., 2008), underscoring the likelihood of a cell-autonomous role for GATA-3 function in T lineage development." . _:b6461379 "S2 B; Sambandam et al., >>2005<<; Lai and Kondo, 2007) and loss of GATA-3 activity affected the development of ETPs during fetal thymopoiesis (Fig." . . _:b6461417 . _:b419252231 . _:b6461341 . _:b419252230 . . _:b6461342 . _:b6461367 . _:b419252225 . _:b6461413 "In fact, production of DN3-like and later stage cells in the presence of few classical ETPs is observed shortly after transplantation (Maillard et al., >>2006<<). A third possibility is the existence of a developmental arrest and/or delay phenotype that causes increased accumulation of Gata3g/g DN3 cells in comparison to Gata3+/+ DN3 cells." . _:b6461343 . _:b419252357 . _:b6461418 . _:b419252320 . _:b419252224 . _:b6461336 . _:b419252193 . . _:b419252227 . _:b6461337 . _:b6461365 "T lineage\u2013restricted progenitors in the fetal liver have been reported (Rodewald et al., 1994; Kawamoto et al., >>1997<<) and have been described within the Lin\u2212c-KithiIL-7R\u03B1+paired immunoglobulin receptor (PIR)+ fraction (Masuda et al., 2005)." . _:b419252226 . . _:b6461338 . _:b6461392 "discussion" . _:b419252237 . _:b419252362 . _:b6461339 . _:b419252236 . _:b6461366 . _:b6461348 . _:b419252235 . _:b419252184 . _:b419252239 . . _:b6461349 . _:b419252238 . _:b6461350 . _:b6461408 "Because Flt3-expressing early ETPs retain B, myeloid, dendritic, and NK cell potential (Porritt et al., 2004; Balciunaite et al., >>2005<<; Sambandam et al., 2005), it is possible that TSP preferentially develops to these non\u2013T lineages in Gata3 mutants." . _:b419252233 . _:b6461351 . _:b419252232 . _:b419252311 . _:b6461344 . . _:b419252365 . _:b6461383 . _:b419252235 . _:b6461345 . _:b6461411 "Similar phenotypes to Gata3g/g hypomorphic mutant embryos are observed in Flt3 ligand\u2013deficient (Sambandam et al., >>2005<<) and E2A null mutant (Dias et al., 2008) mice." . _:b419252350 . _:b419252234 . _:b6461361 . _:b6461346 . _:b6461351 "(lacZ knockin, Gata3z/z; van Doorninck et al., 1999) T cell progenitors in vitro, we isolated and co-cultured fetal liver cells from e12.5/e13.5 catecholamine-rescued Gata3z/z embryos on OP9-DL1 cells (Schmitt and Z\u00FA\u00F1iga-Pfl\u00FCcker, >>2002<<) to promote T cell development. This process was monitored by flow cytometric examination of the cell-surface expression profiles of CD4, CD8, CD44, and CD25." . _:b419252245 . _:b419252229 . _:b6461344 "For fetal liver single-cell preparations, Gata3z/z embryos were recovered from norepinephrine-administered pregnant dams (Kaufman et al., >>2003<<). After PCR genotyping, fetal livers were recovered, minced, and passaged several times through a 5-ml syringe and 26-gauge needle; the single-cell suspension was then subjected to magnetic lineage cell-surface marker depletion before" . . _:b6461347 . _:b419252244 . _:b6461356 . _:b419252247 . _:b6461357 . _:b419252176 . _:b419252246 . _:b6461358 . _:b6461408 . _:b419252241 . _:b6461359 . _:b419252240 . _:b6461352 . _:b419252243 . _:b6461353 . _:b419252242 . _:b6461354 . _:b6461402 "Recently, it was shown that the VCAM-1\u2212 pool of adult LMPPs express GATA-3 mRNA, but GATA-3 in this subfraction is very low when compared with its abundance in ETPs (Lai and Kondo, >>2007<<), so the relevance of this observation is unclear." . _:b419252178 . _:b419252253 . _:b6461355 . _:b419252252 . _:b6461364 . _:b419252251 . _:b419252255 . _:b6461365 . . _:b419252223 . _:b419252306 . _:b419252254 . _:b6461366 . . _:b419252249 . _:b6461367 . . _:b419252248 . _:b6461360 . . _:b6461399 . _:b419252251 . _:b6461361 . . _:b419252250 . _:b6461362 . _:b419252261 . _:b6461363 . _:b419252373 . _:b419252260 . _:b419252313 . _:b6461372 . _:b6461420 "stages, for example in Th2 cells (Amsen et al., 2007; Fang et al., 2007), in contrast, expression of GATA-3 in bone marrow LSK progenitors actually precedes the activation of Notch signaling (Sambandam et al., 2005; Lai and Kondo, >>2007<<), and Notch signaling is dispensable for HSC function in adult mice (Maillard et al., 2008)." . . . _:b419252263 . _:b6461373 . _:b419252159 . _:b419252262 . _:b6461340 . _:b6461374 . _:b419252158 . _:b6461386 "A), retain residual potential for the generation of T cells (for review see Hardy et al., >>2007<<). Because GATA-3 is expressed in these fractions (Akashi et al., 2000; Rumfelt et al., 2006), the developmental potential of Gata3z/z cells in these fractions was examined in adoptive transfer experiments. As shown in Fig. 6 F, the" . _:b419252257 . _:b6461417 "Although examples exist showing that Notch signaling controls GATA-3 expression at other stages, for example in Th2 cells (Amsen et al., >>2007<<; Fang et al., 2007), in contrast, expression of GATA-3 in bone marrow LSK progenitors actually precedes the activation of Notch signaling (Sambandam et al., 2005; Lai and Kondo, 2007), and Notch signaling is dispensable for HSC function" . _:b6461375 . . _:b419252256 . _:b6461368 . . _:b419252259 . _:b6461369 . _:b419252258 . _:b6461370 . _:b6461430 . _:b419252269 . _:b6461371 . _:b419252165 . _:b419252268 . _:b6461380 . _:b419252386 . _:b419252164 . . _:b419252271 . _:b6461381 . _:b419252167 . _:b6461375 "quite heterogeneous in nature, and contain a mixture of mature and immature NK, DC, B, and myeloid lineage cells, as well as rare T cell progenitors (Allman et al., 2003; Porritt et al., 2004; Balciunaite et al., 2005; Anderson, >>2006<<). A recent study also revealed that GATA-3 is expressed in and is required for the generation of thymic NK cells (Vosshenrich et al., 2006)." . _:b419252393 . _:b419252270 . _:b6461399 "LMPPs in the bone marrow are likely to contain some of the cells that acquire a migratory signal that allows thymic homing (Schwarz and Bhandoola, 2004; Adolfsson et al., >>2005<<; Lai and Kondo, 2007; Schwarz et al., 2007)." . _:b6461382 . _:b419252166 . _:b419252265 . _:b6461383 . _:b419252161 . _:b419252264 . _:b6461376 . _:b419252160 . . _:b419252314 . _:b419252267 . _:b6461377 . _:b419252163 . _:b419252266 . . _:b6461378 . _:b419252162 . . _:b419252277 . _:b419252228 . . _:b6461379 . _:b419252173 . _:b419252195 . . _:b419252276 . _:b6461388 . _:b419252172 . _:b6461378 . _:b419252279 . . _:b6461389 . _:b419252175 . . _:b419252278 . _:b6461390 . _:b419252174 . _:b419252218 . _:b419252273 . _:b419252160 . _:b6461391 . _:b419252169 . . _:b419252272 . _:b6461384 . _:b419252168 . _:b419252286 . _:b419252275 . _:b6461385 . _:b419252171 . _:b419252274 . . _:b6461386 . . _:b419252170 . . _:b419252285 . _:b6461387 . _:b419252181 . _:b419252284 . _:b6461396 . _:b419252180 . _:b419252287 . _:b6461397 . _:b419252183 . _:b419252286 . _:b6461398 . _:b419252171 . _:b6461376 "A recent study also revealed that GATA-3 is expressed in and is required for the generation of thymic NK cells (Vosshenrich et al., >>2006<<)." . _:b419252182 . _:b419252281 . . _:b6461399 . _:b419252177 . _:b419252280 . _:b419252176 . _:b6461392 . _:b419252283 . _:b6461393 . _:b419252179 . _:b419252402 . _:b419252282 . _:b6461394 . _:b419252178 . _:b419252293 . _:b6461395 . _:b419252189 . . _:b419252292 . _:b6461404 . _:b419252188 . . . _:b419252295 . _:b6461405 . _:b419252191 . _:b6461365 . _:b419252325 . _:b6461339 . _:b419252294 . _:b6461406 . . _:b419252190 . _:b419252289 . _:b6461407 . _:b419252185 . _:b419252288 . _:b6461400 . _:b419252184 . . _:b419252291 . _:b6461401 . _:b419252187 . _:b6461397 . _:b419252290 . _:b6461402 . _:b419252186 . _:b419252416 . _:b419252379 . _:b419252385 . _:b419252262 . _:b419252301 . _:b419252308 . _:b6461403 . _:b419252197 . . . _:b419252300 . _:b6461412 . _:b419252196 . _:b419252303 . _:b6461413 . _:b419252199 . _:b6461378 "Almost all c-Kithi DN thymocytes, including the earliest T progenitors, do not express NK1.1 at the adult stage (Balciunaite et al., >>2005<<). These results collectively indicated that the contribution of GATA-3 to early T cell progenitor stages would be impossible to assess in the classically defined DN1 (CD4\u2212CD8\u2212CD44+CD25\u2212) cell population, and therefore, we sought to" . _:b419252302 . . _:b6461414 . _:b419252198 . _:b419252297 . _:b6461415 . _:b419252193 . . _:b419252296 . _:b6461408 . _:b419252192 . _:b419252329 . _:b419252299 . _:b6461409 . _:b419252195 . _:b419252298 . _:b419252168 . _:b6461410 . _:b419252194 . _:b419252309 . . _:b6461411 . _:b419252205 . _:b6461337 "Gata3z/z embryos were rescued by norepinephrine administration as previously described (Kaufman et al., >>2003<<). The Gata3flox allele was provided by F. Grosveld (Erasmus Medical Center, Rotterdam, Netherlands; Kurek et al., 2007). C57BL/6-Ly5B6 (CD45.2) mice and C57BL/6-Ly5SJL (CD45.1) mice were purchased from the Jackson Laboratory. All animal" . _:b419252308 . . _:b419252199 . _:b6461420 . . _:b419252204 . _:b419252311 . . _:b6461421 . _:b419252207 . _:b6461362 "Although MPPs have been characterized as the most efficient T cell progenitors at the adult stage (Schwarz and Bhandoola, >>2004<<; Schwarz et al., 2007), the precise characteristics and phenotype of the most efficient prethymic T progenitor cells during fetal development are unknown." . _:b419252310 . _:b6461422 . _:b419252206 . _:b6461374 "cells are actually quite heterogeneous in nature, and contain a mixture of mature and immature NK, DC, B, and myeloid lineage cells, as well as rare T cell progenitors (Allman et al., 2003; Porritt et al., 2004; Balciunaite et al., >>2005<<; Anderson, 2006). A recent study also revealed that GATA-3 is expressed in and is required for the generation of thymic NK cells (Vosshenrich et al., 2006)." . _:b419252305 . _:b6461423 . _:b419252201 . _:b6461404 . _:b419252304 . _:b6461416 . _:b419252200 . . _:b419252307 . _:b6461417 . _:b419252203 . _:b419252256 . _:b419252306 . . _:b6461418 . _:b419252202 . . _:b419252317 . _:b6461419 . _:b419252213 . _:b419252316 . _:b6461428 . _:b419252212 . _:b419252319 . . _:b6461429 . _:b419252215 . _:b419252318 . _:b6461430 . _:b419252214 . _:b419252313 . _:b419252209 . . _:b419252312 . _:b6461424 . _:b419252208 . _:b6461374 . _:b419252315 . _:b6461425 . _:b419252211 . _:b419252314 . . _:b6461426 . _:b6461412 "Similar phenotypes to Gata3g/g hypomorphic mutant embryos are observed in Flt3 ligand\u2013deficient (Sambandam et al., 2005) and E2A null mutant (Dias et al., >>2008<<) mice. There are several possibilities to account for the increased number of Gata3g/g DN2/DN3 stage cells compared with ETPs. First, Gata3 hypomorphic cells may proliferate excessively. In support of this possibility, Gata3z/z" . _:b419252210 . _:b419252325 . _:b6461427 . _:b419252221 . _:b419252324 . _:b419252336 . _:b419252220 . . _:b419252327 . _:b419252223 . _:b419252179 . _:b419252326 . . _:b419252222 . _:b419252321 . _:b419252162 . _:b419252217 . _:b419252320 . _:b419252216 . _:b419252323 . _:b419252254 . _:b419252219 . _:b419252322 . _:b6461347 "results" . _:b419252218 . . _:b419252333 . . _:b6461393 . _:b419252388 . _:b419252332 . _:b419252335 . _:b419252330 . _:b419252334 . . _:b419252329 . _:b419252328 . _:b419252331 . . . _:b419252330 . . . . _:b419252341 . _:b6461390 . _:b419252340 . _:b419252343 . _:b419252367 . . _:b419252342 . _:b419252337 . _:b419252356 . _:b419252336 . _:b419252173 . _:b419252339 . . _:b419252290 . _:b419252338 . . . _:b419252349 . . _:b6461377 . . _:b419252174 . _:b419252348 . _:b419252351 . _:b419252161 . _:b6461360 "The first wave of T progenitor cell migration out of the embryonic fetal liver into the thymic primordium is observed at around e12 to e13 in mice (Owen and Ritter, >>1969<<; Jotereau et al., 1987)." . . _:b419252350 . _:b419252345 . . _:b6461392 . _:b419252344 . . . _:b6461340 "T-restricted progenitors in the fetal liver were analyzed as previously described (Masuda et al., >>2005<<). A cocktail containing antibodies recognizing TER119, Gr-1, B220, CD19, NK1.1, and Thy1.2 was used to exclude Lin+ cells in the fetal liver. Progenitors in the bone marrow were analyzed as previously described (Sambandam et al., 2005;" . _:b6461349 "Although diminished GATA-3 activity in fetal liver cells was reported to result in a loss of Thy-1+ T cells after expansion in fetal thymus organ culture (Hattori et al., 1996; Hozumi et al., >>2008<<), the specific affected stages remained unclear." . _:b419252347 . _:b419252302 . _:b419252346 . _:b419252357 . _:b419252272 . . _:b6461429 "(Sambandam et al., 2005; Schwarz et al., 2007; Sitnicka et al., 2007), IL-7R\u03B1 signaling (Sitnicka et al., 2007), CCR9 activity (Schwarz and Bhandoola, 2004; Svensson et al., 2008), E2A (Dias et al., 2008), Mef2c (Stehling-Sun et al., >>2009<<), and RUNX activity (Talebian et al., 2007) are all known to be important for the generation of ETPs and/or their progenitors, much more extensive characterization will be required to validate the relationships between and epistatic" . _:b419252356 . _:b6461339 "Immature T cells were analyzed as previously described (Talebian et al., >>2007<<). The following cocktail of antibodies was used to exclude mature hematopoietic lineage cells (Lin+) in the thymus and the cells cultured in the OP9-DL co-culture system:" . _:b419252359 . _:b419252358 . _:b419252392 . _:b419252353 . _:b419252339 . . _:b419252352 . _:b419252337 . _:b419252355 . _:b6461405 . . _:b419252232 . _:b419252167 . _:b419252354 . _:b419252188 . _:b419252365 . _:b419252364 . _:b6461347 . _:b419252367 . . _:b6461363 . _:b419252366 . _:b419252361 . _:b419252360 . _:b6461334 . _:b419252363 . _:b419252362 . . _:b419252261 . _:b6461396 "In this paper, we show that the vast majority of GATA-3\u2013expressing cells in the DN1 compartment also expressed NK1.1, which is abundantly expressed on c-Kit\u2212 cells of the CD4\u2212CD8\u2212CD44+ population (Laurent et al., >>2004<<; Balciunaite et al., 2005)." . _:b419252373 . _:b419252401 . _:b419252194 . _:b419252372 . _:b419252375 . _:b419252374 . . _:b419252369 . . _:b419252368 . . _:b419252219 . . _:b419252371 . _:b419252192 . . _:b6461364 "T lineage\u2013restricted progenitors in the fetal liver have been reported (Rodewald et al., >>1994<<; Kawamoto et al., 1997) and have been described within the Lin\u2212c-KithiIL-7R\u03B1+paired immunoglobulin receptor (PIR)+ fraction (Masuda et al., 2005)." . _:b419252370 . _:b419252381 . _:b419252380 . _:b6461376 . . _:b419252383 . _:b419252307 . . _:b419252382 . _:b419252291 . . _:b419252377 . _:b6461406 "A current candidate for the TSPs resides in the pool of ETPs expressing both Flt3 and CCR9 (Benz et al., >>2008<<). Because the generation of ETPs was almost completely blocked in mice that harbored a conditionally inactivated Gata3 gene at the adult stage, we speculate that GATA-3 activity is required at this step. Because Flt3-expressing early ETPs" . _:b419252376 . . _:b419252379 . . _:b419252378 . _:b419252389 . _:b419252214 . _:b6461353 "Gata3 null mutant mice fail to generate even a rudimentary thymus (Lim et al., 2000; Moriguchi et al., >>2006<<), whereas Gata3 conditionally mutant mice that were crossed to mice bearing an lck-directed Cre transgene, and therefore lacked Gata3 at the DN3 stage and thereafter, were reported to have normal numbers of DN-stage cells per thymus (DN1" . _:b419252388 . _:b419252391 . . _:b419252406 . _:b419252390 . _:b419252186 . _:b419252248 . . . _:b419252385 . _:b419252384 . _:b6461403 "Introduction of GATA-3 before Notch signaling activation fails to promote T cell development (Hozumi et al., >>2008<<). In the absence of GATA-3, bone marrow immature NK cells develop normally, whereas thymic NK cells are not generated (Samson et al., 2003; Vosshenrich et al., 2006). These data suggest a function for GATA-3 in a lymphocyte progenitor" . . _:b419252387 . _:b419252386 . _:b419252397 . _:b419252349 . _:b419252303 . _:b419252396 . _:b6461414 "Perhaps because of significance within this context, Notch signaling has been documented to be required for the development of ETPs (Sambandam et al., >>2005<<). Both GATA-3 and Notch signaling are required for T cell development, whereas either factor alone cannot induce thymocyte development (Hozumi et al., 2008). When Lin\u2212c-Kit+ fetal liver cells are cultured on OP9-DL1 cells, Notch target" . _:b6461372 . _:b419252399 . . _:b6461372 "more recent studies have revealed that DN1 cells are actually quite heterogeneous in nature, and contain a mixture of mature and immature NK, DC, B, and myeloid lineage cells, as well as rare T cell progenitors (Allman et al., >>2003<<; Porritt et al., 2004; Balciunaite et al., 2005; Anderson, 2006). A recent study also revealed that GATA-3 is expressed in and is required for the generation of thymic NK cells (Vosshenrich et al., 2006)." . _:b6461338 . _:b419252398 . _:b419252163 . _:b419252393 . _:b419252292 . _:b419252392 . _:b419252328 . _:b419252395 . . _:b419252394 . _:b419252405 . _:b419252333 . _:b6461406 . _:b419252404 . _:b419252407 . _:b419252406 . _:b419252401 . _:b6461346 "It should be noted that the B220 epitope is expressed on early to mature B lineage cells as well as on some activated T, dendritic, and NK cells (Watanabe and Akaike, 1994; Huntington et al., >>2007<<)." . _:b419252169 . _:b419252293 . _:b419252400 . _:b6461386 . _:b6461345 . _:b419252403 . _:b419252402 . _:b419252370 . _:b419252413 . _:b419252412 . _:b419252415 . _:b6461336 "Yamamoto, Tohoku University Medical School, Sendai, Japan; K\u00FChn et al., >>1995<<) have been previously described." . _:b419252414 . _:b419252175 . _:b419252295 . . _:b419252377 . _:b419252409 . _:b6461351 . _:b419252368 . . . _:b419252408 . . _:b419252411 . _:b6461387 "Because GATA-3 is expressed in these fractions (Akashi et al., >>2000<<; Rumfelt et al., 2006), the developmental potential of Gata3z/z cells in these fractions was examined in adoptive transfer experiments." . _:b419252410 . _:b419252201 . . . . . _:b6461371 . _:b6461336 . _:b419252417 . _:b419252416 . _:b419252408 . _:b419252418 . _:b6461380 "S2 B; Sambandam et al., 2005; Lai and Kondo, >>2007<<) and loss of GATA-3 activity affected the development of ETPs during fetal thymopoiesis (Fig." . _:b419252415 . _:b6461373 . _:b419252237 . _:b419252381 . _:b419252403 . _:b419252206 . _:b419252364 . _:b419252264 . _:b419252205 . _:b419252326 . . _:b6461423 .