Pluripotency Genes Overexpressed in Primate Embryonic Stem Cells Are Localized on Homologues of Human 16, 17, 19 and X

While human embryonic stem cells (hESCs) are predisposed towards chromosomal aneploidities on 12, 17, 20 and X, rendering them susceptible to transformation, the specific genes expressed are not yet known. Here, by identifying the genes over expressed in pluripotent rhesus ESCs (nhpESCs) and comparing them to both their genetically-identical differentiated progeny (teratoma fibroblasts) as well as genetically-related differentiated parental cells (parental skin fibroblasts from whom gametes were used for ESC derivation), we find that some of those over expressed genes in nhpESCs cluster preferentially on rhesus chromosomes 16, 19, 20 and X, homologues of human chromosomes 17, 19, 16 and X respectively. Differentiated parental skin fibroblasts display gene expression profiles closer to nhpESC profiles than to teratoma cells, which are genetically identical to the pluripotent nhpESCs. Twenty over and under expressed pluripotency modulators, some implicated in neurogenesis, have been identified. The over expression of some of these genes discovered using pedigreed nhpESCs derived from prime embryos generated by fertile primates, which is impossible to perform with the anonymously donated clinically-discarded embryos from which hESCs are derived, independently confirms the importance of chromosome 17 and X regions in pluripotency and suggests specific candidates for targeting differentiation and transformation decisions. To overcome the genetic background heterogeneity which might mask some aspects of the actual stemness signature, we utilized a newly established bank of pedigreed non-human primate embryonic stem cells (nhpESCs) to generate a unique non-human primate “stemness gene” list generated by the comparison of nhpESC gene expression to two different sources of genetically-identical or genetically-related differentiated fibroblasts. The first were grown out of skin explants taken from the nonhuman primate parents from which gametes were used to generate the stem cell lines. The second were teratoma explants. RNA was isolated from all three types of cells: 1. Skin fibroblasts; these samples are indicated by the letter M (monkey) before the monkey identification number and sample label (a-f). 2. nhpESCs denoted by C (cell line) followed by the cell line and sample. 3. Fibroblast explants from teratomas generated by the injection of nhpESC into immune compromised mice. Since there were a number of teratomas for every stem cell line, we designated the nomenclature as follows: T (teratoma) - number of teratoma from the line - line name - sample. Therefore, TA31b denotes that this is the second sample of the first teratoma from line 3106. A total of 39 samples were used for this comparison: Pluripotent pedigreed rhesus ESCs (female ESC line C3806; male ESC line C3106) were compared to genetically identical differentiated progeny grown from their resultant teratomas (five male teratomas and three female teratomas: ‘T’, i.e. TA31, TB31, TC31, TD31, TE31 and TA38, TB38, TC38 respectively) as well as to differentiated cells from their macaque parents (‘M’). therefore, 2 cell lines (3106 and 3806) 3 types of skin fibroblasts and 8 teratoma fibroblasts (5 generated from line 3106 and 3 from 3806). All samples were run in biological triplicates for a total of 39 samples. A total of 39 samples were used for this comparison: Pluripotent pedigreed rhesus ESCs (female ESC line C3806; male ESC line C3106) were compared to genetically identical differentiated progeny grown from their resultant teratomas (five male teratomas and three female teratomas: ‘T’, i.e. TA31, TB31, TC31, TD31, TE31 and TA38, TB38, TC38 respectively) as well as to differentiated cells from their macaque parents (‘M’). therefore, 2 cell lines (3106 and 3806) 3 types of skin fibroblasts and 8 teratoma fibroblasts (5 generated from line 3106 and 3 from 3806). All samples were run in biological triplicates for a total of 39 samples.