Transcriptional coregulator Ess2 controls survival of post-thymic CD4+ T cells through the Myc and IL-7 signaling pathways

Abstract

Ess2, also known as Dgcr14, is a transcriptional co-regulator of CD4+ T cells. Ess2 is located in a chromosomal region, the loss of which has been associated with 22q11.2 deletion syndrome (22q11DS), which causes heart defects, skeletal abnormalities, and immunodeficiency. However, the specific association of Ess2 with 22q11DS remains unclear. To elucidate the role of Ess2 in T-cell development, we generated Ess2 floxed (Ess2fl/fl) and CD4+ T cell–specific Ess2 KO (Ess2ΔCD4/ΔCD4) mice using the Cre/loxP system. Interestingly, Ess2ΔCD4/ΔCD4 mice exhibited reduced naïve T-cell numbers in the spleen, while the number of thymocytes (CD4−CD8−, CD4+CD8+, CD4+CD8−, and CD4−CD8+) in the thymus remained unchanged. Furthermore, Ess2ΔCD4/ΔCD4 mice had decreased NKT cells and increased γδT cells in the thymus and spleen. A genome-wide expression analysis using RNA-seq revealed that Ess2 deletion alters the expression of many genes in CD4 single-positive thymocytes, including genes related to the immune system and Myc target genes. In addition, Ess2 enhanced the transcriptional activity of c-Myc. Some genes identified as Ess2 targets in mice show expressional correlation with ESS2 in human immune cells. Moreover, Ess2ΔCD4/ΔCD4 naïve CD4+ T cells did not maintain survival in response to IL-7. Our results suggest that Ess2 plays a critical role in post-thymic T-cell survival through the Myc and IL-7 signaling pathways. Ess2, also known as Dgcr14, is a transcriptional co-regulator of CD4+ T cells. Ess2 is located in a chromosomal region, the loss of which has been associated with 22q11.2 deletion syndrome (22q11DS), which causes heart defects, skeletal abnormalities, and immunodeficiency. However, the specific association of Ess2 with 22q11DS remains unclear. To elucidate the role of Ess2 in T-cell development, we generated Ess2 floxed (Ess2fl/fl) and CD4+ T cell–specific Ess2 KO (Ess2ΔCD4/ΔCD4) mice using the Cre/loxP system. Interestingly, Ess2ΔCD4/ΔCD4 mice exhibited reduced naïve T-cell numbers in the spleen, while the number of thymocytes (CD4−CD8−, CD4+CD8+, CD4+CD8−, and CD4−CD8+) in the thymus remained unchanged. Furthermore, Ess2ΔCD4/ΔCD4 mice had decreased NKT cells and increased γδT cells in the thymus and spleen. A genome-wide expression analysis using RNA-seq revealed that Ess2 deletion alters the expression of many genes in CD4 single-positive thymocytes, including genes related to the immune system and Myc target genes. In addition, Ess2 enhanced the transcriptional activity of c-Myc. Some genes identified as Ess2 targets in mice show expressional correlation with ESS2 in human immune cells. Moreover, Ess2ΔCD4/ΔCD4 naïve CD4+ T cells did not maintain survival in response to IL-7. Our results suggest that Ess2 plays a critical role in post-thymic T-cell survival through the Myc and IL-7 signaling pathways. Transcriptional co-regulators contribute to gene expression by interacting with DNA-bound transcription factors (1Rosenfeld M.G. Lunyak V.V. Glass C.K. Sensors and signals: a coactivator/corepressor/epigenetic code for integrating signal-dependent programs of transcriptional response.Genes Dev. 2006; 20: 1405-1428Crossref PubMed Scopus (777) Google Scholar). The functions of transcription co-regulators are diverse, including histone modifications, chromatin structure conversion, and chromosome structure changes. Because they regulate multiple transcription factors, transcriptional co-regulator functions in vivo are complex and still under investigation. We recently identified Ess2 (also known as Dgcr14 or Es2) as a transcriptional co-regulator of retinoid-related orphan nuclear receptor gamma/gamma-t (RORγ/γt) through biochemical purification and matrix-assisted laser desorption ionization–time of flight mass spectrometry analysis (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar). In humans, the ESS2 gene was first cloned as an expression sequence tag located in the 22q11.2 locus, which is related to the 22q11.2 deletion syndrome (22q11DS; also known as DiGeorge syndrome or CATCH 22 syndrome) (3Lindsay E.A. Rizzu P. Antonacci R. Jurecic V. Delmas-Mata J. Lee C.C. et al.A transcription map in the CATCH22 critical region: identification, mapping, and ordering of four novel transcripts expressed in heart.Genomics. 1996; 32: 104-112Crossref PubMed Scopus (42) Google Scholar, 4Rizzu P. Lindsay E.A. Taylor C. O'Donnell H. Levy A. Scambler P. et al.Cloning and comparative mapping of a gene from the commonly deleted region of DiGeorge and Velocardiofacial syndromes conserved in C. elegans.Mamm. Genome. 1996; 7: 639-643Crossref PubMed Scopus (24) Google Scholar). 22q11DS is a disorder caused by a small deletion, located near the middle of chromosome 22, specifically designated q11.2 (5Scambler P.J. The 22q11 deletion syndromes.Hum. Mol. Genet. 2000; 9: 2421-2426Crossref PubMed Scopus (407) Google Scholar, 6Robin N.H. Shprintzen R.J. Defining the clinical spectrum of deletion 22q11.2.J. Pediatr. 2005; 147: 90-96Abstract Full Text Full Text PDF PubMed Scopus (182) Google Scholar, 7McDonald-McGinn D.M. Sullivan K.E. Chromosome 22q11.2 deletion syndrome (DiGeorge syndrome/Velocardiofacial syndrome).Medicine (Baltimore). 2011; 90: 1-18Crossref PubMed Scopus (343) Google Scholar). Common signs and symptoms of this syndrome include heart abnormalities, cleft palate, distinctive facial features, and schizophrenia. In addition, 22q11DS patients are often immunodeficient and experience recurrent infections, and some patients develop autoimmune disorders, including rheumatoid arthritis and Graves’ disease (8Sullivan K.E. Chromosome 22q11.2 deletion syndrome and DiGeorge syndrome.Immunol. Rev. 2019; 287: 186-201Crossref PubMed Scopus (73) Google Scholar). While genetically modified mouse models expressing 22q11DS phenotypes have been reported, immunodeficiency-related genes within 22q11.2 have not yet been identified. RORγ/γt are members of the nuclear receptor superfamily, and they regulate TH17 cell development (9Mangelsdorf D.J. Thummel C. Beato M. Herrlich P. Schutz G. Umesono K. et al.The nuclear receptor superfamily: the second decade.Cell. 1995; 83: 835-839Abstract Full Text PDF PubMed Scopus (6146) Google Scholar, 10Jetten A.M. Retinoid-related orphan receptors (RORs): critical roles in development, immunity, circadian rhythm, and cellular metabolism.Nucl. Recept. Signal. 2009; 7e003Crossref PubMed Scopus (512) Google Scholar). Ess2 regulates Il17a mRNA levels, which are induced during TH17 cell development through RORγ/γt (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar). Additionally, the genes ribosomal S6 kinase 2 (Rsk2; Rps6ka3) and bromodomain adjacent to zinc finger domain 1B (Baz1b) associate with Ess2 to regulate the transcriptional activities of RORγ/γt in TH17 cells (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar). Ess2 mRNA expression levels are higher in TH17 cells than in naïve, TH1, TH2, or induced regulatory T cells (iTregs) (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar). The Ess2 protein localizes in the nucleus where it interacts with both transcription factors and spliceosomes (11Bessonov S. Anokhina M. Will C.L. Urlaub H. Luhrmann R. Isolation of an active step I spliceosome and composition of its RNP core.Nature. 2008; 452: 846-850Crossref PubMed Scopus (304) Google Scholar, 12Hegele A. Kamburov A. Grossmann A. Sourlis C. Wowro S. Weimann M. et al.Dynamic protein-protein interaction wiring of the human spliceosome.Mol. Cell. 2012; 45: 567-580Abstract Full Text Full Text PDF PubMed Scopus (282) Google Scholar, 13Takada I. Tsuchiya M. Yanaka K. Hidano S. Takahashi S. Kobayashi T. et al.Ess2 bridges transcriptional regulators and spliceosomal complexes via distinct interacting domains.Biochem. Biophys. Res. Commun. 2018; 497: 597-604Crossref PubMed Scopus (11) Google Scholar). The function of Ess2 during T-cell development in vivo is poorly understood because homozygous Ess2 mutant mice are embryonically lethal (14Lindsay E.A. Botta A. Jurecic V. Carattini-Rivera S. Cheah Y.C. Rosenblatt H.M. et al.Congenital heart disease in mice deficient for the DiGeorge syndrome region.Nature. 1999; 401: 379-383Crossref PubMed Google Scholar). Our previous studies (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar) suggested the possibility that Ess2 regulates T-cell development and differentiation in vivo. To elucidate the function of the Ess2 gene during T-cell development in vivo, we generated Ess2 gene floxed mice (Ess2fl/fl) and CD4-specific Ess2 knockout (KO) mice (Ess2ΔCD4/ΔCD4) by crossing CD4-Cre transgenic mice (15Lee P.P. Fitzpatrick D.R. Beard C. Jessup H.K. Lehar S. Makar K.W. et al.A critical role for Dnmt1 and DNA methylation in T cell development, function, and survival.Immunity. 2001; 15: 763-774Abstract Full Text Full Text PDF PubMed Scopus (974) Google Scholar) and Ess2fl/fl mice. Interestingly, these mice survive but have reduced naïve T-cell numbers. Our studies on these mice have elucidated the role of the Ess2 gene during T-cell development. To elucidate Ess2 function in vivo, we generated Ess2ΔCD4/ΔCD4 mice using the Cre-loxP system (Fig. S1A). We first generated Ess2 KO (Ess2−/−) and Ess2+/− mice by inserting a splicing acceptor into the exon 2, FRP, and loxP sequences (Fig. S1A). Consistent with previous reports, Ess2−/− mice were embryonically lethal (Fig. S1, B and C) (14Lindsay E.A. Botta A. Jurecic V. Carattini-Rivera S. Cheah Y.C. Rosenblatt H.M. et al.Congenital heart disease in mice deficient for the DiGeorge syndrome region.Nature. 1999; 401: 379-383Crossref PubMed Google Scholar). However, Ess2+/− mice survived and did not have any abnormal appearances or bodyweight loss (data not shown). We then crossed Ess2+/− mice with FLP-expressing transgenic mice (RIKEN No. RBRC01834, see Experimental procedures) to generate Ess2fl/fl mice and To the role of Ess2 during T-cell development, we generated Ess2ΔCD4/ΔCD4 by crossing Ess2fl/fl with CD4-Cre transgenic mice (15Lee P.P. Fitzpatrick D.R. Beard C. Jessup H.K. Lehar S. Makar K.W. et al.A critical role for Dnmt1 and DNA methylation in T cell development, function, and survival.Immunity. 2001; 15: 763-774Abstract Full Text Full Text PDF PubMed Scopus (974) Google Ess2fl/fl were as We the of the Ess2 in CD4 single-positive and thymocytes from Ess2ΔCD4/ΔCD4 mice by the of a (Fig. Ess2ΔCD4/ΔCD4 mice and have a to Ess2fl/fl mice (Fig. Ess2 mRNA expression levels were reduced in thymocytes from Ess2ΔCD4/ΔCD4 mice but not in thymocytes (Fig. results that the Ess2 gene in Ess2ΔCD4/ΔCD4 mice was deleted in thymocytes and deleted in We the number of T cells in the of Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice. were in the spleen, and the number of was in both Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice (Fig. However, the numbers of (Fig. 2, B and and T cells (Fig. 2, were in the of Ess2ΔCD4/ΔCD4 mice to Ess2fl/fl mice. The of CD4+ T cells to and T cells to were both decreased in Ess2ΔCD4/ΔCD4 mice (Fig. 2, and The of CD4+ T cells to T cells was reduced in Ess2ΔCD4/ΔCD4 the of T cells to T cells was not (Fig. 2, and In addition, the number of naïve CD4+ T cells was of in Ess2fl/fl mice (Fig. 2, and While the of naïve CD4+ T cells to was also in Ess2ΔCD4/ΔCD4 the of naïve CD4+ T cells to CD4+ T cells was to the Ess2fl/fl mice (Fig. However, the naïve the of Ess2ΔCD4/ΔCD4 CD4+ T cells to T cells or CD4+ T cells was than that of the Ess2fl/fl mice (Fig. Interestingly, Ess2ΔCD4/ΔCD4 mice had γδT cells and T cells than Ess2fl/fl mice (Fig. 2, and was in regulatory T cells (Fig. naïve CD4+ T cells were under TH17 (Fig. the mRNA expression levels of genes and were in the Ess2ΔCD4/ΔCD4 mice. suggest that the loss of the Ess2 gene alters T-cell development in the of Ess2ΔCD4/ΔCD4 mice. Our results that Ess2 regulates naïve T-cell development To Ess2 is active in T-cell development, we the number of and thymocytes in the of Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice. Ess2 mRNA expression levels in and thymocytes did not Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice (Fig. results suggest that the deletion of Ess2 in Ess2ΔCD4/ΔCD4 mice during the and were in both mouse (Fig. and in Ess2ΔCD4/ΔCD4 mice B and and We the expression of regulatory genes and in thymocytes by (Fig. The gene regulates the of thymocytes from the thymus C.C. CD4+ T cells in of deletion by of or PubMed Scopus Google Scholar, T. S. K. H. et are for of PubMed Scopus Google and is in T-cell development to the of T cells from the thymus M. G. V. et from thymus and is on receptor PubMed Scopus Google Scholar). and mRNA expression levels did not Ess2ΔCD4/ΔCD4 and Ess2fl/fl mice (Fig. γδT cells (Fig. also while NKT cells decreased (Fig. in the thymus of Ess2ΔCD4/ΔCD4 mice. results that Ess2 not contribute to the of T cells or from the However, Ess2 the development of γδT cells and NKT cells in the Our results that an Ess2 deletion in thymocytes naïve T-cell the is poorly To this we the expression of CD4+ genes in thymocytes and naïve T cells from Ess2fl/fl and we did not any (Fig. To elucidate the role of Ess2 in naïve T-cell we the genome-wide expression of and thymocytes as as naïve CD4 T-cell using RNA-seq and A and we many in mRNA expression in thymocytes but a in thymocytes (Fig. A and a splicing were in Ess2ΔCD4/ΔCD4 mice (data not shown). previous studies show that Ess2 interacts with both the spliceosome and transcription factors I. Tsuchiya M. Yanaka K. Hidano S. Takahashi S. Kobayashi T. et al.Ess2 bridges transcriptional regulators and spliceosomal complexes via distinct interacting domains.Biochem. Biophys. Res. Commun. 2018; 497: 597-604Crossref PubMed Scopus (11) Google results that Ess2 gene transcription the of development. However, the of genes expressed Ess2ΔCD4/ΔCD4 and Ess2fl/fl mice in and naïve T cells by to (Fig. results suggest that Ess2 regulates gene in cells than in naïve T cells. We the in expression of genes in genes had than a in expression and some genes were expressed in naïve T cells but not cells (Fig. To the clinical of ESS2 we the and Interestingly, genes were with ESS2 expression in the human disease of CD4+ and T cells from patients et from patients are for and gene Scholar) results suggest that they are in the of the immune system in and that ESS2 expression a in autoimmune we a for and analysis and a gene analysis The analysis revealed that genes in and cell and I were in both Ess2ΔCD4/ΔCD4 thymocytes and naïve T cells to Ess2fl/fl cells (Fig. results show that Ess2 regulates genes from multiple immune pathways. We the gene with a the Myc and were also in Ess2ΔCD4/ΔCD4 thymocytes and naïve T cells (Fig. In Ess2ΔCD4/ΔCD4 thymocytes, and gene were were also in Ess2ΔCD4/ΔCD4 mice (Fig. and Myc as an transcription in T and R. S. R. et al.The transcription Myc T 2011; 35: Full Text Full Text PDF PubMed Scopus Google Scholar). Interestingly, a an interaction Myc and Ess2 through analysis M. C. et protein distinct that to Cell. 2018; Full Text Full Text PDF PubMed Scopus Google Scholar). on these we that Ess2 as a transcriptional co-regulator of To elucidate the association Ess2 and a was by Ess2 and with a that in the cell in Ess2 enhanced the transcriptional activity of in a Additionally, chromatin analysis that naïve CD4+ T cells from Ess2ΔCD4/ΔCD4 mice the of Ess2 to the which a H. C. G. et of for and S. A. PubMed Scopus Google Scholar). naïve CD4+ T cells from Ess2ΔCD4/ΔCD4 mice did not the of to the (Fig. the levels of the active histone histone decreased in these cells. Moreover, of and in cells revealed in the nucleus (Fig. results that Ess2 has a on the transcriptional activity of c-Myc. we the and a correlation ESS2 expression and the genes identified by the specifically in of with S. V. C. V. C. et expression of cells from cell into its cellular and to 2009; PubMed Scopus Google and B cells and disease R. I. H. et of an in 2008; PubMed Scopus Google and A and ESS2 expression levels with some of the genes in human results show that ESS2 have functions in and which is through gene The IL-7 signaling regulates naïve T-cell survival and P. J. M. T. M. et of T 2000; PubMed Scopus Google Scholar, C.L. The many of from to T cell 2005; PubMed Scopus Google and mRNA was reduced in cells from Ess2ΔCD4/ΔCD4 mice A and B and we that Ess2 regulate CD4+ T-cell analysis that deletion of the Ess2 gene decreased the of the histone to the (Fig. We naïve CD4+ T cells from Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice with or IL-7 for IL-7 the survival of cells from Ess2fl/fl but not from Ess2ΔCD4/ΔCD4 (Fig. We a number of Ess2ΔCD4/ΔCD4 naïve CD4+ T cells of the IL-7 (Fig. and with IL-7 to T-cell we did not in the gene expression of Ess2ΔCD4/ΔCD4 naïve CD4+ T cells (Fig. we cell by with and IL-7. in Ess2ΔCD4/ΔCD4 naïve CD4+ T cells reduced results suggest that Ess2 regulates naïve T-cell survival through the IL-7 signaling In this we generated Ess2fl/fl mice and CD4-specific Ess2 KO mice for the first (14Lindsay E.A. Botta A. Jurecic V. Carattini-Rivera S. Cheah Y.C. Rosenblatt H.M. et al.Congenital heart disease in mice deficient for the DiGeorge syndrome region.Nature. 1999; 401: 379-383Crossref PubMed Google we that Ess2−/− mice had an (Fig. results that Ess2 plays an role in the is unclear. Ess2ΔCD4/ΔCD4 mice reduced T-cell and numbers but had increased numbers of γδT cells. In Ess2ΔCD4/ΔCD4 the of CD4+ T cells to or T cells Additionally, the of naïve CD4+ T cells to CD4+ T cells while the of CD4+ T cells to T cells or CD4+ T cells in the of naïve and CD4+ T cells in Ess2ΔCD4/ΔCD4 mice to the that ESS2 has on cell survival in naïve or CD4+ T cells. studies in to this expression revealed that Ess2 regulates genes in cells. In the expression of Myc target genes and genes was decreased in Ess2ΔCD4/ΔCD4 thymocytes and naïve T which the of reduced naïve T-cell in these mice. Interestingly, that Ess2 as a transcriptional co-regulator for the transcriptional activity of and that Ess2 is with in the In addition, the expression levels of Myc target as and are with ESS2 expression in immunodeficient patients (Fig. results suggest that Ess2 CD4+ T-cell survival and via transcriptional through and RORγ/γt (Fig. studies elucidate the transcriptional show that deletion of Ess2 caused transcriptional than splicing suggest a role for Ess2 as a transcriptional co-regulator in the immune system. Additionally, show that Ess2ΔCD4/ΔCD4 mice in both γδT and NKT cells. γδT cells develop from thymocytes in the and some transcription factors that regulate development have been identified thymus to of cell Rev. PubMed Scopus Google Scholar). is CD4-Cre deficient mice increased the number of and γδT cells S. A. K. H. S. K. et receptor development and of of S. A. PubMed Scopus Google Scholar). Because Ess2ΔCD4/ΔCD4 naïve T cells exhibited reduced mRNA levels (Fig. A and as as IL-7 signal-dependent cell survival (Fig. we that of the IL-7 signaling to maintain is Ess2 regulates the function of and transcription factors in the IL-7 signaling Furthermore, are many transcription factors in differentiation thymus to of cell Rev. PubMed Scopus Google and it is which of these are by studies on the function of Ess2 in γδT cells. NKT cells are a of T that are and distinct from CD4+ and T cells R. K.E. Transcriptional of NKT cell Rev. PubMed Scopus Google Scholar). While T cells and NKT cells both in the are development in the thymus from a of thymocytes and transcription factors, as and Because Ess2 localizes to the nucleus and regulates RORγ/γt NKT cells in Ess2ΔCD4/ΔCD4 mice RORγ/γt activity in studies are to elucidate the role of Ess2 on the transcriptional of NKT cells. Interestingly, human ESS2 expression with the expression of and genes in autoimmune disease patients and results suggest that Ess2 also contribute to and in However, a analysis is and a analysis of Ess2 in a KO mouse elucidate the of immune Our previous studies that Ess2 induces TH17 differentiation by the transcriptional activities of RORγ/γt (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar). In Ess2ΔCD4/ΔCD4 gene mRNA levels were also in CD4+ T cells under TH17 (Fig. However, in this we that Ess2 of T-cell which are of transgenic mice to the function of cells in vivo. KO mice regulate naïve T-cell through expression A. S. et for in survival and 2000; PubMed Scopus Google which did not in Ess2ΔCD4/ΔCD4 mice (data not shown). results show that the Ess2 protein with transcription factors to regulate which also contribute to immune mouse models of 22q11DS A. mapping of the 22q11.2 deletion region and the of 2015; 7: PubMed Scopus Google including 22q11.2 deletion mice and transgenic mice A. et a mouse 22q11.2 deletion and of 2015; PubMed Scopus Google Scholar, A. K. K. T. et al.A mouse of 22q11.2 and of disease and 2018; PubMed Scopus Google Scholar, In the deleted genes associated with deletion are they 2019; PubMed Scopus Google Scholar). A has the of a deletion of 22q11DS R. M. T. K. J. et analysis of a novel mouse of the 22q11.2 deletion a with the deletion the human 22q11.2 PubMed Scopus (24) Google Scholar). 22q11DS phenotypes using these mouse as development, abnormal and schizophrenia. However, the genes within the 22q11.2 deletion region that are related to have not yet been We the of from the of Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice under were (data not shown). Our results suggest that a analysis of Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice into 22q11DS as it immunodeficiency. In we generated both Ess2fl/fl and Ess2ΔCD4/ΔCD4 mice and have a role for Ess2 in T-cell to elucidate the role of Ess2 in the of the immune system. To generate Ess2+/− we the Ess2 from (Fig. S1A). The was and into cell from the mouse by and Ess2 cells were cell were then into from mice. were into the of and the mice crossed with mice. Ess2+/− mice were identified in the the of the Ess2 Ess2+/− mice were then for with mice to Ess2+/− mice. We generated the Ess2−/− mice by crossing and Ess2+/− mice. The deletion of Ess2 was by (Fig. To generate Ess2fl/fl mice (Fig. Ess2+/− mice were crossed with transgenic mice No. H. H. S. mice in 2006; PubMed Scopus Google Ess2fl/fl mice were then crossed with CD4-Cre transgenic mice (15Lee P.P. Fitzpatrick D.R. Beard C. Jessup H.K. Lehar S. Makar K.W. et al.A critical role for Dnmt1 and DNA methylation in T cell development, function, and survival.Immunity. 2001; 15: 763-774Abstract Full Text Full Text PDF PubMed Scopus (974) Google and and mice were crossed to generate the Ess2ΔCD4/ΔCD4 mice. Ess2ΔCD4/ΔCD4 mice were with using (Fig. and and (Fig. mice were in specific were to and and and were by specific for CD4 or and were from and an Ess2 specific was generated in I. Tsuchiya M. Yanaka K. Hidano S. Takahashi S. Kobayashi T. et al.Ess2 bridges transcriptional regulators and spliceosomal complexes via distinct interacting domains.Biochem. Biophys. Res. Commun. 2018; 497: 597-604Crossref PubMed Scopus (11) Google and an specific for was from and thymocytes were from to mice and to cell and to generate a were in 2 in with were on a and using was using was from using an or system was with the I The is expressed as where is the the of for and that of the sequences were (2Takada I. DGCR14 induces Il17a gene expression through the RORgamma/BAZ1B/RSKS2 complex.Mol. Cell. Biol. 2015; 35: 344-355Crossref PubMed Scopus (9) Google Scholar) and are CD4 single-positive and naïve T cells were with a cell and was with RNA-seq was using the We from and were first using the RNA-seq were to the mouse from using the an for of novel Biol. 2011; Scopus Google The for gene were using the expression of transcripts was from RNA-seq using to of exon gene expression C. A. G. D.R. et gene and expression analysis of RNA-seq with and 2012; 7: PubMed Scopus Google Scholar). Some were and using To were generated and into H. M. G. et 2011; PubMed Scopus Google of were using using the RNA-seq were using RNA-seq were and gene and were using the gene gene were into the in and analysis of gene using 2009; PubMed Scopus Google Scholar, the analysis of gene Res. 2009; PubMed Scopus Google Scholar). was with the and the for was was using the and gene were from the CD4+ T cells from were using the CD4+ Isolation and in with or and cells were identified using and by and cells were using cell naïve T cells were with a and was and using the TH17 cell naïve T cells were under and 2 for and then the was the cells were using the to the R. A. H. S. K. et of a novel on in Res. 2018; PubMed Scopus Google Scholar). the cells were and for and activity using a and a of the of the expression of the expression and of the Ess2 expression for a of the were to with the of an for in a were to the and results are as the were in and in A was to the cells were with and chromatin was by to and was with or specific and then with protein A for an 2 and were by DNA was by using the system with The was expressed as where is the the for of the and that of the The sequences are in are as the of was using an the RNA-seq were using a or the were or than were an than was The of RNA-seq are on the No. Ess2+/− mice and Ess2fl/fl mice were with the are within this and the The that they have of with the of the We the members of for and M. for with RNA-seq and We the of for with I. T. and S. T. I. S. K. H. M. and A. I. S. S. and S. I. T. S. S. and H. S. S. and H. T. T. S. and M. M. M. M. and was by the and a for and

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