Human Dystrophin Dp71ab Enhances the Proliferation of Myoblasts Across Species But Not Human Nonmyoblast Cells

Abstract

Dystrophin Dp71 is an isoform produced from the Dp71 promoter in intron 62 of the DMD gene, mutations in which cause Duchenne muscular dystrophy. Dp71 is involved in various cellular processes and comprises more than 10 isoforms produced by alternative splicing. Dp71ab, in which both exons 71 and 78 are deleted, has a hydrophobic C-terminus that is hydrophilic in Dp71. Therefore, Dp71ab is believed to have different roles from Dp71. Previously, we reported that Dp71ab enhanced the proliferation of human myoblasts. Here, we further characterized Dp71ab, focusing on the activation of cell proliferation. Dp71ab increased the proliferation of immortalized human myoblasts in a dose-dependent manner. In contrast, Dp71 suppressed proliferation in a dose-dependent manner. Consistent with these opposite effects, eGFP-tagged Dp71ab and mCherry-tagged Dp71 showed different cellular distributions, with Dp71ab mostly in the nucleus. Notably, human Dp71ab enhanced the proliferation of rat and mouse myoblasts. Despite these findings, human Dp71ab did not enhance the proliferation of human nonmyoblast cells, including rhabdomyosarcoma cells. We concluded that Dp71ab is a myoblast-specific proliferation enhancer. In further studies, Dp71ab will be employed for the expansion of myoblasts in clinical settings.

CRISPR-Cas9 correction in the DMD mouse model is accompanied by upregulation of Dp71f protein

“…Over 10 Dp71 splice variants in normal and malignant tissues or model systems have been described. 30 , 33 , 34 Some isoforms are common in undifferentiated satellite cells rather than in myotubes, 35 , 36 and specific amino acids in different splice forms can regulate protein localization. 37 To specify splice variants upregulated in CRISPR-Cas9-treated DMDdel8–34 mice, we amplified full-length Dp71 cDNA (∼2 kb; Figure S8 A) and identified exon content by Sanger sequencing.…”

Nuclear Small Dystrophin Isoforms during Muscle Differentiation

“…Instead, phosphorylation is thought to be the regulator of nuclear import [ 29 ]. It has been shown before that Dp71 enhances the proliferation of myoblasts [ 13 ] and can localize to the nucleus and to the nuclear envelope, where it stabilizes nuclear dystrophin-associated proteins [ 15 , 17 ]. We could confirm the nuclear and nuclear-envelope localization of Dp71.…”

“…The carboxy-terminal isoform Dp71, which is expressed ubiquitously, has been linked to myoblast proliferation [ 13 ] and neural stem/progenitor cell proliferation and differentiation [ 14 ]. This isoform has been shown to localize within and closely around the nucleus of PC12, HeLa, C2C12 and HEK293 cells and human myoblasts [ 13 , 15 , 16 , 17 , 18 , 19 ], but also to mitochondria [ 20 ].…”

“…The carboxy-terminal isoform Dp71, which is expressed ubiquitously, has been linked to myoblast proliferation [ 13 ] and neural stem/progenitor cell proliferation and differentiation [ 14 ]. This isoform has been shown to localize within and closely around the nucleus of PC12, HeLa, C2C12 and HEK293 cells and human myoblasts [ 13 , 15 , 16 , 17 , 18 , 19 ], but also to mitochondria [ 20 ]. Depletion of Dp71 alters the distribution of aquaporin-4 channels in brain macroglial cells, but the question of which function Dp71 fulfils in the nucleus and how stable it appears there remains unclear.…”

“…Life 2023, 13,1367 Conflicts of Interest: The authors declare no conflict of interest. The funders had no role in the study's design; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.…”