ABSTRACT
The differentiation of primordial germ cells (PGCs) is a fundamental step in development. PR domain-containing protein 14 (PRDM14) and B lymphocyte-induced maturation protein 1 (BLIMP1) play pivotal roles in mouse PGC specification. In the present study, we assessed the roles of chicken orthologs of PRDM14 and BLIMP1 in PGC development. PRDM14 and BLIMP1 were expressed in blastodermal cells and PGCs. The in vivo knockdown of PRDM14 or BLIMP1 by introducing a replication-competent retroviral vector expressing shRNAs to the blastodermal stage of embryos reduced the number of SSEA-1 or chicken vasa homologue-positive PGCs on day 5.5-6.5. Since the inhibition of Activin receptor-like kinase 4/5/7 in cultured PGCs reduced the expression of PRDM14, BLIMP1, and NANOG, and that of MEK inhibited PRDM14 expression, the expression of these genes seems to be controlled by Activin A and FGF2 signaling. Overall, PRDM14, BLIMP1, and NANOG seem to be involved in the self-renewal of PGCs in cultured PGCs and embryos.
Subject(s)
Avian Proteins/genetics , Cell Differentiation/genetics , Gene Expression Regulation, Developmental , Germ Cells/metabolism , Positive Regulatory Domain I-Binding Factor 1/genetics , Animals , Avian Proteins/metabolism , Blastoderm/cytology , Blastoderm/metabolism , Cell Self Renewal/genetics , Cells, Cultured , Chick Embryo , Chickens , Germ Cells/cytology , Lewis X Antigen/genetics , Lewis X Antigen/metabolism , Nanog Homeobox Protein/genetics , Nanog Homeobox Protein/metabolism , Positive Regulatory Domain I-Binding Factor 1/metabolism , RNA InterferenceABSTRACT
Ten-eleven translocation (TET) methylcytosine dioxygenase has potential as an active eraser to regulate the genomic DNA methylation status. We herein cloned chicken TET (cTET) family genes, and confirmed their functions. Quantitative reverse-transcription PCR showed that cTET1 was strongly expressed in erythrocytes throughout development. This cTET1 expression pattern, together with the results of methylated or hydroxymethylated DNA immunoprecipitation, suggests that cTET1 contributes to demethylation around the promoter region of the definitive-type ß-globin gene ßΑ in erythroid cells. The knockdown of cTET1 in T2ECs chicken erythroid progenitor cells suppressed the induction of ßΑ expression under differentiation conditions. These results suggest that cTET1 plays an important role in erythroid cell differentiation.
Subject(s)
5-Methylcytosine/metabolism , Chickens/genetics , Cloning, Molecular , Dioxygenases/genetics , Erythropoiesis , 5-Methylcytosine/analogs & derivatives , Animals , Cell Line , Chickens/physiology , DNA Methylation , Dioxygenases/metabolism , HeLa Cells , Humans , Multigene Family , Promoter Regions, Genetic , beta-Globins/geneticsABSTRACT
In mammals, interferon-inducible transmembrane proteins (IFITMs) prevent infections by various enveloped viruses. The expression of IFITMs in chicken was herein examined in the adult and embryonic organs using a quantitative reverse-transcription-polymerase chain reaction. The results obtained revealed that IFITM3 was expressed at a higher level than IFITM1, 2 and 5, in both embryonic and adult organs. However, the expression levels of IFITMs in embryonic organs were less than 5 % of those in adult lungs. Among the embryonic tissues examined, primordial germ cells (PGCs) at day 2.5 expressed relatively higher levels of IFITM3. IFITM3 expression levels were 1.5-fold higher in the chicken cell line DF-1 than in PGCs. The knockdown of IFITM3 in DF-1 cells by siRNA increased the infectivity of a vesicular stomatitis virus G protein-pseudotyped lentiviral vector, suggesting that lower levels of IFITM3 are still sufficient to restrict this viral vector.