Independent regulation of vertebral number and vertebral identity by microRNA-196 paralogs

Abstract

The Hox genes play a central role in patterning the embryonic anterior-to-posterior axis. An important function of Hox activity in vertebrates is the specification of different vertebral morphologies, with an additional role in axis elongation emerging. The miR-196 family of microRNAs (miRNAs) are predicted to extensively target Hox 3′ UTRs, although the full extent to which miR-196 regulates Hox expression dynamics and influences mammalian development remains to be elucidated. Here we used an extensive allelic series of mouse knockouts to show that the miR-196 family of miRNAs is essential both for properly patterning vertebral identity at different axial levels and for modulating the total number of vertebrae. All three miR-196 paralogs, 196a1, 196a2, and 196b, act redundantly to pattern the midthoracic region, whereas 196a2 and 196b have an additive role in controlling the number of rib-bearing vertebra and positioning of the sacrum. Independent of this, 196a1, 196a2, and 196b act redundantly to constrain total vertebral number. Loss of miR-196 leads to a collective up-regulation of numerous trunk Hox target genes with a concomitant delay in activation of caudal Hox genes, which are proposed to signal the end of axis extension. Additionally, we identified altered molecular signatures associated with the Wnt, Fgf, and Notch/segmentation pathways and demonstrate that miR-196 has the potential to regulate Wnt activity by multiple mechanisms. By feeding into, and thereby integrating, multiple genetic networks controlling vertebral number and identity, miR-196 is a critical player defining axial formulae.

MicroRNA-196a2 Biomarker and Targetome Network Analysis in Solid Tumors

“…These results are consistent with previous studies reporting that upregulation of miR-196a was found to be correlated with advanced tumor stage and poor overall and recurrence-free survival in many cancer patients [8,52]. This upregulation could suppress annexin A1; an inhibitor of cell proliferation, and mediator of apoptosis and anchorage-independent growth [68], enhance G1/S-phase transition and the proliferative ability of cancer cells through targeting FOXO1 and p27Kip1; two key effectors of the PI3K/Akt signaling pathway that regulates cell proliferation and has been addressed as a therapeutic target [8,9], up-regulate the ubiquitin-conjugating enzyme E2C (UBE2C) proto-oncogene [83], inhibit Bach1 (a basic leucine zipper mammalian transcriptional repressor) and upregulate hemeoxygenase 1 in HCV-related HCC [84], inhibit IjBa, which binds and tethers NF-jB in the cytoplasm; leading to translocation of the later into the nucleus where it activates a variety of target genes such as apoptosis-related genes Bax and Bcl-2 that regulate activation of caspase-3 [65], and regulates the Wnt-Fgf-Notch signaling pathways [85] that are commonly associated with tissue invasion and metastasis [86]. Together, these findings suggest that miR-196a2 could be a favorable prognostic biomarker in a cancer-specific pattern.…”

MicroRNA filters Hox temporal transcription noise to confer boundary formation in the spinal cord

“…For example, miR-10 resides in almost all taxa between Hox4 and 5 paralogs and arose in early bilaterians, while miR-196 is located between Hox9 and 10 paralogs and is specific to vertebrates and urochordates. Genetic knockout or overexpression studies further indicate that Hox-embedded miRNAs are involved in regulating Hox gene expression at the post-transcriptional level1516171819. Interestingly, while Hox genes are transcribed in spinal progenitors, many Hox proteins are only detectable in postmitotic MNs720.…”

Hoxb6 can interfere with somitogenesis in the posterior embryo through a mechanism independent of its rib-promoting activity

“…These included miR196, which is known to be involved in the regulation of several Hox genes (Yekta et al, 2004(Yekta et al, , 2008. In addition, deletion of the three miR196 family members produced alterations in the identity and/or number of vertebrae in the mouse (Wong et al, 2015). This area of the Hoxb6 3′UTR also contains a target for miR199, mutants of which display skeletal defects (Watanabe et al, 2008).…”

“…In favor of this hypothesis, the Hoxb6 3′UTR contains several potential miR binding sites that are conserved between mouse and human. Particularly interesting are those for miR199, as its inactivation results in malformed vertebrae (Watanabe et al, 2008), and those for the miR196 family because mice lacking all three members of this family have extra lumbar ribs (Wong et al, 2015), resembling the effect of ectopic Hoxb6 activation. However, removing the binding sites for these two miRs from the Hoxb6 3′UTR had no effect on the development of the axial skeleton or on Hoxb6 expression.…”