Fine-tuning of Hh signaling by the RNA-binding protein Quaking to control muscle development

Abstract

SUMMARYThe development of the different muscles within the somite is a complex process that involves the Hedgehog (Hh) signaling pathway. To specify the proper number of muscle cells and organize them spatially and temporally, the Hh signaling pathway needs to be precisely regulated at different levels, but only a few factors external to the pathway have been described. Here, we report for the first time the role of the STAR family RNA-binding protein Quaking A (QkA) in somite muscle development. We show in zebrafish that the loss of QkA function affects fast muscle fiber maturation as well as Hh-induced muscle derivative specification and/or morphogenesis. Mosaic analysis reveals that fast fiber maturation depends on the activity of QkA in the environment of fast fiber progenitors. We further show that Hh signaling requires QkA activity for muscle development. By an in silico approach, we screened the 3ЈUTRs of known Hh signaling component mRNAs for the Quaking response element and found the transcription factor Gli2a, a known regulator of muscle fate development. Using destabilized GFP as a reporter, we show that the gli2a mRNA 3ЈUTR is a functional QkA target. Consistent with this notion, the loss of QkA function rescued slow muscle fibers in yot mutant embryos, which express a dominant-negative Gli2a isoform. Thus, our results reveal a new mechanism to ensure muscle cell fate diversity by fine-tuning of the Hh signaling pathway via RNA-binding proteins.

Structure–function studies of STAR family Quaking proteins bound to their in vivo RNA target sites

“…Substitution of the hydrophobic Val side chain pointing toward the core of the KH domain for a polar Glu is likely to abrogate hydrophobic contacts with Lys167 and Leu168 of a6 and C119 at the beginning of the b2 strand and thereby destabilize the overall fold of the KH domain. Recently, another mutation in the hydrophobic b3/a6 interface in zebrafish quaking protein has been found to lead to severe developmental defects (Lobbardi et al 2011) by changing a conserved Ile (Ile155 in QKI) (Fig. 3E) to Asn with destabilizing effects possibly similar to those of the mouse V157E mutation.…”

“…Thus, mRNAs of the myelin basic protein (MBP) (Larocque et al 2002), the oligodendrocyte cell differentiation factor CDKN1B/p27(Kip1) (Larocque et al 2005), and the alternative splicing regulator HNRNPA1 (Zearfoss et al 2011) in humans as well as the muscle-specific transcription factor Gli2a in zebrafish (Lobbardi et al 2011) are stabilized as a result of specific interactions of QKI with specific sequence elements in their 39 untranslated regions (UTRs). While QR proteins show high evolutionary conservation on the sequence level, their regulatory mechanisms are very different, as, in the case of GLD-1, binding of target mRNAs leads to translational repression in vitro and in vivo rather than mRNA stabilization (Lee and Schedl 2010).…”

Quaking and PTB control overlapping splicing regulatory networks during muscle cell differentiation

“…Quaking has been implicated in a striking variety of processes in the mouse, such as embryogenesis, blood vessel development, glial cell fate determination, apoptosis, and smooth muscle development, while the human homolog, QKI, has been implicated in a number of diseases, including ataxia, glioblastoma development, and schizophrenia (Chénard and Richard 2008). QK homologs are expressed in genomes as divergent as sea urchin and are critically involved in fundamental cell and developmental processes in deeply diverged metazoans from C. elegans (gld-1, asd-2) (Francis et al 1995;Ohno et al 2008) to Drosophila (how) (Baehrecke 1997;Fyrberg et al 1997;Zaffran et al 1997) and zebrafish (qkA) (Tanaka et al 1997;Lobbardi et al 2011). In vertebrates, a larger family of proteins related to QK also exists, including SAM68 and Overlapping Quaking and PTB splicing networks www.rnajournal.org 633 the less-studied SLM-1 and SLM-2 (Lock et al 1996;Di Fruscio et al 1999).…”

Expression analysis of Gli1 and Gli2 in different tissues and muscle-derived cells of Qinchuan cattle

“…Although the expression levels of Gli1 and Gli2 in muscle are both the lowest of the 13 tissues we investigated, many previous researches have testified that Hh signaling is crucial in muscle formation during embryonic and postnatal development of vertebrates (Flynt et al, 2007;Martin et al, 2007;Straface et al, 2009;Lobbardi et al, 2011;Hu et al, 2012). At the cellular level, the Hh signaling pathway promotes the proliferation and myogenic differentiation of myoblasts (Pola et al, 2003;Li et al, 2004;Koleva et al, 2005;Elia et al, 2007;Madhala-Levy et al, 2012).…”

“…Additionally, the repression of the Hh signaling pathway induced by cyclopamine or a small interfering RNA (siRNA) of its positive component inhibited chondrogenic differentiation (Wu et al, 2013). The Hh signaling pathway promotes myogenesis in zebrafish (Flynt et al, 2007;Lobbardi et al, 2011), Xenopus (Martin et al, 2007), mouse (Straface et al, 2009;Hu et al, 2012;Voronova et al, 2013), and chick (Elia et al, 2007).…”