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Background: The differentiation of human bone marrow derived skeletal stem cells (known as human bone marrow stromal or mesenchymal stem cells, hMSCs) into osteoblasts involves the activation of a small number of well-described transcription factors. To identify additional osteoblastic transcription factors, we studied gene expression of hMSCs during ex vivo osteoblast differentiation., Results: Clustering of gene expression, and literature investigation, revealed three transcription factors of interest - ZNF25, ZNF608 and ZBTB38. siRNA knockdown of ZNF25 resulted in significant suppression of alkaline phosphatase (ALP) activity. This effect was not present for ZNF608 and ZBTB38. To identify possible target genes of ZNF25, we analyzed gene expression following ZNF25 siRNA knockdown. This revealed a 23-fold upregulation of matrix metallopeptidase 1 and an 18-fold upregulation of leucine-rich repeat containing G protein-coupled receptor 5 and RAN-binding protein 3-like. We also observed enrichment in extracellular matrix organization, skeletal system development and regulation of ossification in the entire upregulated set of genes. Consistent with its function as a transcription factor during osteoblast differentiation of hMSC, we showed that the ZNF25 protein exhibits nuclear localization and is expressed in osteoblastic and osteocytic cells in vivo. ZNF25 is conserved in tetrapod vertebrates and contains a KRAB (Krueppel-associated box) transcriptional repressor domain., Conclusions: This study shows that the uncharacterized transcription factor, ZNF25, is associated with differentiation of hMSC to osteoblasts.

Krüppel-like factors 3 and 8 (KLF3 and KLF8) are highly related transcriptional regulators that bind to similar sequences of DNA. We have previously shown that in erythroid cells there is a regulatory hierarchy within the KLF family, whereby KLF1 drives the expression of both the Klf3 and Klf8 genes and KLF3 in turn represses Klf8 expression. While the erythroid roles of KLF1 and KLF3 have been explored, the contribution of KLF8 to this regulatory network has been unknown. To investigate this, we have generated a mouse model with disrupted KLF8 expression. While these mice are viable, albeit with a reduced life span, mice lacking both KLF3 and KLF8 die at around embryonic day 14.5 (E14.5), indicative of a genetic interaction between these two factors. In the fetal liver, Klf3 Klf8 double mutant embryos exhibit greater dysregulation of gene expression than either of the two single mutants. In particular, we observe derepression of embryonic, but not adult, globin expression. Taken together, these results suggest that KLF3 and KLF8 have overlapping roles in vivo and participate in the silencing of embryonic globin expression during development.

Abstract: The hippocampus and cerebellum represent anatomically and functionally distinct parts of the human brain. The RNA-Seq technique makes it possible to investigate the human transcriptome with unprecedented resolution, allowing identification of differential mRNA splicing and promoter usage on a genome-wide scale. We undertook whole-mRNA sequencing of samples from the human hippocampus and cerebellum. A bioinformatic analysis revealed distinct expression patterns of genes related to the molecular physiology of neurons and glial cells. Upregulated genes in hippocampal tissue included serpin peptidase inhibitor, clade A (SERPINA3), lymphocyte antigen 6 complex, locus H (LY6H) and transthyretin (TTR). In cerebellum, the cerebellin 3 precursor (CLBN3) and Zic family member 4 (ZIC4) genes were significantly upregulated. These changes were validated in independent donor samples by qRT-PCR. The hippocampus and the cerebellum showed striking differences in splicing patterns and promoter usage. A notable example of this was the gene for NGFI-A binding protein 2 (NAB2), which displayed tissue-specific isoforms which may affect its function as a transcriptional repressor. [Copyright &y& Elsevier]