NF-YA, the regulatory subunit of the trimeric transcription factor (TF) NF-Y, is regulated by alternative splicing (AS) generating two major isoforms, "long" (NF-YAl) and "short" (NF-YAs). Muscle cells express NF-YAl. We ablated exon 3 in mouse C2C12 cells by a four-guide CRISPR/Cas9n strategy, obtaining clones expressing exclusively NF-YAs (C2-YAl-KO). C2-YAl-KO cells grow normally, but are unable to differentiate. Myogenin and-to a lesser extent, MyoD- levels are substantially lower in C2-YAl-KO, before and after differentiation. Expression of the fusogenic Myomaker and Myomixer genes, crucial for the early phases of the process, is not induced. Myomaker and Myomixer promoters are bound by MyoD and Myogenin, and Myogenin overexpression induces their expression in C2-YAl-KO. NF-Y inactivation reduces MyoD and Myogenin, but not directly: the Myogenin promoter is CCAAT-less, and the canonical CCAAT of the MyoD promoter is not bound by NF-Y in vivo. We propose that NF-YAl, but not NF-YAs, maintains muscle commitment by indirectly regulating Myogenin and MyoD expression in C2C12 cells. These experiments are the first genetic evidence that the two NF-YA isoforms have functionally distinct roles.
The Switch from NF-YAl to NF-YAs Isoform Impairs Myotubes Formation / D. Libetti, A. Bernardini, S. Sertic, G. Messina, D. Dolfini, R. Mantovani. - In: CELLS. - ISSN 2073-4409. - 9:3(2020 Mar), pp. 789.1-789.21.
|Titolo:||The Switch from NF-YAl to NF-YAs Isoform Impairs Myotubes Formation|
MANTOVANI, ROBERTO (Corresponding)
|Parole Chiave:||C2C12 cells; CRISPR-Cas9; NF-Y; exon deletion; muscle differentiation; splicing isoforms.|
|Settore Scientifico Disciplinare:||Settore BIO/17 - Istologia|
Settore BIO/18 - Genetica
Settore BIO/11 - Biologia Molecolare
|Data di pubblicazione:||mar-2020|
|Digital Object Identifier (DOI):||http://dx.doi.org/10.3390/cells9030789|
|Appare nelle tipologie:||01 - Articolo su periodico|