Abstract
BackgroundIK is a splicing factor that promotes spliceosome activation and contributes to pre-mRNA splicing. Although the molecular mechanism of IK has been previously reported in vitro, the physiological role of IK has not been fully understood in any animal model. Here, we generate an ik knock-out (KO) zebrafish using the CRISPR/Cas9 system to investigate the physiological roles of IK in vivo.ResultsThe ik KO embryos display severe pleiotropic phenotypes, implying an essential role of IK in embryonic development in vertebrates. RNA-seq analysis reveals downregulation of genes involved in skeletal muscle differentiation in ik KO embryos, and there exist genes having improper pre-mRNA splicing among downregulated genes. The ik KO embryos display impaired neuromuscular junction (NMJ) and fast-twitch muscle development. Depletion of ik reduces myod1 expression and upregulates pax7a, preventing normal fast muscle development in a non-cell-autonomous manner. Moreover, when differentiation is induced in IK-depleted C2C12 myoblasts, myoblasts show a reduced ability to form myotubes. However, inhibition of IK does not influence either muscle cell proliferation or apoptosis in zebrafish and C2C12 cells.ConclusionThis study provides that the splicing factor IK contributes to normal skeletal muscle development in vivo and myogenic differentiation in vitro.
Highlights
IK is a splicing factor that promotes spliceosome activation and contributes to pre-mRNA splicing
CRISPR/Cas9-mediated ik KO embryos display abnormal phenotypes and lethality Zebrafish contains a single copy of ik (Genebank Accession number BC049322.1) on chromosome 21; the encoded protein is 548 amino acids in length
These results indicate that IK plays an essential role in zebrafish embryonic development
Summary
IK is a splicing factor that promotes spliceosome activation and contributes to pre-mRNA splicing. Pre-mRNA splicing is performed by a sequential assembly of five small nuclear RNA–proteins (snRNPs) called the spliceosome [1,2,3,4]. IK, known as RED protein because of its arginine (R)-, glutamic acid (E)-, and aspartic acid (D)-rich domain, participates in the regulation of cell mitotic kinases and phosphatases [10] and localization of the spindle assembly checkpoint protein MAD1 to the kinetochore [11]. It plays a role in pre-mRNA splicing by interacting with SMU1, a B complex-specific protein. To elucidate the functional role of IK during development, we established, for the first time, the ik knock-out (KO) zebrafish model with the CRISPR/Cas system
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
