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There is ample evidence that the ubiquitin-proteasome system is an important regulator of transcription and its activity is necessary for maintaining pluripotency and promoting cellular reprogramming. Moreover, proteasome activity contributes to maintaining the open chromatin structure found in pluripotent stem cells, acting as a transcriptional inhibitor at specific gene loci generally associated with differentiation. The current study was designed to understand further the role of proteasome inhibition in reprogramming and its ability to modulate endogenous expression of pluripotency-related genes and induced pluripotent stem cells (iPSCs) colony formation. Herein, we demonstrate that acute combinatorial treatment with the proteasome inhibitors MG101 or MG132 and the histone deacetylase (HDAC) inhibitor valproic acid (VPA) increases gene expression of the pluripotency marker Oct3/4, and that MG101 alone is as effective as VPA in the induction of Oct3/4 mRNA expression in fibroblasts. Prolonged proteasome inhibition cyclically upregulates gene expression of Oct3/4 and Nanog, but reduces colony formation in the presence of the iPSC induction cocktail. In conclusion, our results demonstrate that the 26S proteasome is an essential modulator in the reprogramming process. Its inhibition enhances expression of pluripotency-related genes; however, efficient colony formation requires proteasome activity. Therefore, discovery of small molecules that increase proteasome activity might lead to more efficient cell reprogramming and generation of pluripotent cells. [ABSTRACT FROM AUTHOR]

Hyperplasia is a major contributor to the increase in adipose tissue mass that is characteristic of obesity. However, the identity and characteristics of cells that can be committed into adipocyte lineage remain unclear. Stem cell antigen 1 (Sca-1) has been used recently as a candidate marker in the search for tissue-resident stem cells. In our quest for biomarkers of cells that can become adipocytes, we analyzed ear mesenchymal stem cells (EMSC), which can differentiate into adipocytes, osteocytes, chondrocytes, and myocytes. Our previous studies have demonstrated that EMSC abundantly expressed Sca-1. In the present study, we have analyzed the expression of adipogenic transcription factors and adipocyte-specific genes in Sca-1-enriched and Sca-1-depleted EMSC fractions. Sca- 1-enriched EMSC accumulated more lipid droplets during adipogenic differentiation than Sca-1-depleted. Similarly, EMSC isolated from Sca-1-/- mice displayed reduced lipid accumulation relative to EMSC from wildtype controls (p < .01). Comparative analysis of the adipogenic differentiation process between Sca-1-enriched and Sca-1-depleted populations of EMSC revealed substantial differences in the gene expression. Preadipocyte factor 1, CCAAT enhancer-binding protein (C/EBP) β, C/EBPα, peroxisome proliferator-activated receptor γ2, lipoprotein lipase, and adipocyte fatty acid binding protein were expressed at significantly higher levels in the Sca-1-enriched EMSC fraction. However, the most striking observation was that leptin was detected only in the conditioned medium of Sca-1-enriched EMSC. In addition, we performed loss-of-function (Sca-1 morpholino oligonucleotide) experiments. The data presented here suggest that Sca-1 is a biomarker for EMSC with the potential to become functionally active adipocytes. [ABSTRACT FROM AUTHOR]