miR-30 family controls proliferation and differentiation of intestinal epithelial cell models by directing a broad gene expression program that includes SOX9 and the ubiquitin ligase pathway

Bailey C E Peck, John Sincavage, Sydney Feinstein, Amanda T. Mah, James G. Simmons, P. Kay Lund, Praveen Sethupathy

Research output: Research - peer-reviewArticle

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Abstract

Proliferation and differentiation of intestinal epithelial cells (IECs) occur in part through precise regulation of key transcription factors, such as SOX9. MicroRNAs (miRNAs) have emerged as prominent fine-tuners of transcription factor expression and activity. We hypothesized that miRNAs, in part through the regulation of SOX9, may mediate IEC homeostasis. Bioinformatic analyses of the SOX9 3′-UTR revealed highly conserved target sites for nine different miRNAs. Of these, only the miR-30 family members were both robustly and variably expressed across functionally distinct cell types of the murine jejunal epithelium. Inhibition of miR-30 using complementary locked nucleic acids (LNA30bcd) in both human IECs and human colorectal adenocarcinoma-derived Caco-2 cells resulted in significant up-regulation of SOX9 mRNA but, interestingly, significant down-regulation of SOX9 protein. To gain mechanistic insight into this non-intuitive finding, we performed RNA sequencing on LNA30bcd-treated human IECs and found 2440 significantly increased genes and 2651 significantly decreased genes across three time points. The up-regulated genes are highly enriched for both predicted miR-30 targets, as well as genes in the ubiquitin-proteasome pathway. Chemical suppression of the proteasome rescued the effect of LNA30bcd on SOX9 protein levels, indicating that the regulation of SOX9 protein by miR-30 is largely indirect through the proteasome pathway. Inhibition of the miR-30 family led to significantly reduced IEC proliferation and a dramatic increase in markers of enterocyte differentiation. This in-depth analysis of a complex miRNA regulatory program in intestinal epithelial cell models provides novel evidence that the miR-30 family likely plays an important role in IEC homeostasis.

LanguageEnglish (US)
Pages15975-15984
Number of pages10
JournalJournal of Biological Chemistry
Volume291
Issue number31
DOIs
StatePublished - Jul 29 2016

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Ligases
Ubiquitin
Epithelial Cells
Gene Expression
Gene expression
MicroRNAs
Genes
Proteasome Endopeptidase Complex
Proteins
Homeostasis
Transcription Factors
Inhibition (Psychology)
RNA Sequence Analysis
Caco-2 Cells
Enterocytes
Differentiation Antigens
3' Untranslated Regions
Computational Biology
Adenocarcinoma
Up-Regulation

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

miR-30 family controls proliferation and differentiation of intestinal epithelial cell models by directing a broad gene expression program that includes SOX9 and the ubiquitin ligase pathway. / Peck, Bailey C E; Sincavage, John; Feinstein, Sydney; Mah, Amanda T.; Simmons, James G.; Lund, P. Kay; Sethupathy, Praveen.

In: Journal of Biological Chemistry, Vol. 291, No. 31, 29.07.2016, p. 15975-15984.

Research output: Research - peer-reviewArticle

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