Novel Variants of Oct-3/4 Gene Expressed in Mouse Somatic Cells

Abstract

It has been suggested that Oct-3/4 may regulate self-renewal in somatic stem cells, as it does in embryonic stem cells. However, recent reports raise the possibility that detection of human Oct-3/4 expression by RT-PCR is prone to artifacts generated by pseudogene transcripts and argue against a role for Oct-3/4 in somatic cells. In this study, we clarified Oct-3/4 expression in mouse somatic tissues using designed PCR primers, which can exclude amplification of its pseudogenes. We found that novel alternative transcripts are indeed expressed in somatic tissues, rather than the normal length transcripts in germline and ES cells. The alternative transcripts indicate the expression of two kinds of truncated proteins. Furthermore, we determined novel promoter regions that are sufficient for the expression of Oct-3/4 transcript variants in somatic cells. These findings provide new insights into the postnatal role of Oct-3/4 in somatic tissues.Oct-3/4 is one of the earliest transcription factors expressed in the embryo, and its orthologs have been found in many different animal species. The nuclear protein is encoded by a homeobox-containing gene Pou5f1, which belongs to the family of POU (Pit Oct Unc) genes (1-3). It regulates gene transcription by binding DNA to octamer motifs ATGCA/TAAT. For many years, Oct-3/4 has been recognized as a gatekeeper for maintaining pluripotency in embryonic stem (ES) 2 cells and pre-implantation embryos. Pluripotent state of ES cells is influenced by expression levels of Oct-3/4 gene (4, 5). More recently, it was demonstrated that mature adult cells could be "reprogrammed" to a very primitive embryonic state via forced expression of four genes (Oct-3/4, c-Myc, Klf-4, and Sox-2) (6); Oct-3/4 is an essential reprogramming factor in this case.Another line of studies have shown Oct-3/4 expression in somatic stem cells and cancer cells (refer to supplement data in Ref. 7). In mouse epithelial tissues, ectopic expression of Oct-3/4 blocks progenitor cell differentiation and causes dysplasia (8). Another report suggests that Oct-3/4 plays a critical role in the genesis of germ cell tumors (9). In addition, Oct-3/4 expression is reported in adult normal tissues (e.g. hematopoietic and mesenchymal stem cells, pancreatic islets, kidney, brain, etc.). These results suggest that Oct-3/4 may not be only crucial for maintaining pluripotency in pre-implantation embryos and germline cells.Although many investigators have reported Oct-3/4 expression in a number of somatic stem cells and cancer cells, this remains a controversial and unsolved issue because of the existence of Oct-3/4 pseudogenes. A number of Oct-3/4 pseudogenes have been identified in humans, and some Oct-3/4 pseudogenes have also been suggested in mouse (10 -14). Many of the results obtained from previous studies for Oct-3/4 expression in somatic tissues using RT-PCR analysis were confusing because PCR primer sets used in the analysis were unable to clearly distinguish Oct-3/4 from its pseudogenes. In fact, Cantz et al. (14) have r...

Identification and differential expression patterns of porcine OCT4 variants

“…OCT4B expression was confirmed in human pancreatic islet (Takeda et al 1992) and mouse eye, especially in retinal pigment epithelial cells (Mizuno & Kosaka 2008). The expression of OCT4B in somatic tissue indicates that the variant could have a different function compared with OCT4A and would have little relationship with pluripotency.…”

“…The structure of variant mRNA is similar between human and mice, including POU-and CTD-coding exons with differential NTD-coding exon compared with the A form. However, the sequence of the OCT4B-specific exon and NTDs were not conserved between human and mice (Mizuno & Kosaka 2008, Guo et al 2012. According to our result, the porcine genome aligned to the human OCT4B-specific exon with low homology (Fig.…”

“…These studies have made OCT4A an essential gene for analyzing stem cell status and embryo development in various species, including pig. However, compared with the OCT4A, other variants have been less focused even though several years have passed since the identification of the variant in human and mice (Takeda et al 1992, Mizuno & Kosaka 2008. The OCT4 variants have been focused with regard to the following two aspects generally: i) distinct properties compared with OCT4A (Cauffman et al 2006, Lee et al 2006a, Atlasi et al 2008, Guo et al 2012 and ii) an inducer making the improper OCT4A detection (Wang & Dai 2010).…”

“…The OCT4B transcript shares identical exons coding the POU DNA-binding and C-terminal transactivation domains (CTDs) of OCT4A, but the variants have different exon-coding N-terminal transactivation domains (NTDs) (Takeda et al 1992, Mizuno & Kosaka 2008. Moreover, OCT4B1, which is composed of the same exons with OCT4B, contains an additional cryptic exon, which is an intron region in OCT4A and OCT4B transcripts (Atlasi et al 2008, Wang & Dai 2010.…”

“…Human and mouse OCT4 variants, OCT4B and OCT4B1, have been identified (Takeda et al 1992, Atlasi et al 2008, Mizuno & Kosaka 2008 and their expressions have been examined in preimplantation embryos, ES cells, EC cells, somatic cells, and various cancer cells (Cauffman et al 2006, Lee et al 2006a, Atlasi et al 2008, Mizuno & Kosaka 2008, Guo et al 2012. The OCT4B transcript shares identical exons coding the POU DNA-binding and C-terminal transactivation domains (CTDs) of OCT4A, but the variants have different exon-coding N-terminal transactivation domains (NTDs) (Takeda et al 1992, Mizuno & Kosaka 2008.…”