A Novel Dnmt3a1 Transcript Inhibits Adipogenesis

Abstract

DNA (cytosine-5)-methyltransferase 3a (Dnmt3a) is an enzyme that catalyzes the transfer of methyl groups to specific CpG forms in DNA. In mammals, two variant transcripts of Dnmt3a have been successfully identified. To the best of our knowledge, no Dnmt3a transcripts in an avian have been successfully identified. This study was performed to detect different transcripts of Dnmt3a in chickens and to examine whether a novel Dnmt3a transcript named Dnmt3a1 may regulate adipogenesis. In addition to cloning, sequencing, transcript detection, and expression studies, a novel Dnmt3a1 transcript overexpression and knockdown were conducted to explore the potential role of Dnmt3a1 in preadipocyte proliferation and the early stage of adipocyte differentiation. In chicken abdominal fat tissue, we detected a novel Dnmt3a1 transcript that differs from Dnmt3a by lacking 23 amino acids at the exon-1/exon-2 border. Dnmt3a1 mRNA was ubiquitously expressed in a variety of tissues or cells and highly expressed in chicken adipose tissue/cells. The expression of Dnmt3a1 was regulated under different physiological conditions including aging, fasting, and high-fat diet. In addition, overexpression of Dnmt3a1 significantly decreased preadipocyte proliferation and induced cell-cycle arrest while its inhibition increased cell proliferation and S-phase cells. Furthermore, the overexpression of Dnmt3a1 significantly upregulated the mRNA level of cell-cycle-related genes, such as CDKN1A, CDKN1B, CCNB3, CCND2, CCNG2, CDKN2B, and CDK9, or the protein level of CDKN1A, CDKN1B, and CCNG2. Conversely, the knockdown of Dnmt3a1 by siRNA had the opposite effects. Moreover, during early adipocyte differentiation, the overexpression of Dnmt3a1 significantly decreased the mRNA and the protein levels of PPAR-γ, C/EBP-α, ADIPOR1, and STAT3, and the mRNA levels of FAS, LEPR, LPL, PRKAB2, and ATGL. In contrast, their expression was significantly increased after the knockdown of Dnmt3a1. Taken together, we identified a novel transcript of Dnmt3a, and it played a potential role in adipogenesis.

The Landscape of DNA Methylation Associated With the Transcriptomic Network of Intramuscular Adipocytes Generates Insight Into Intramuscular Fat Deposition in Chicken

“…Growing number of studies suggested that DNA methylation played significantly role in adipogenesis (Broholm et al, 2016;Chen et al, 2016;Lim et al, 2016). Previous studies recommended that DNMT3A inhibited porcine intramuscular preadipocytes differentiation by changing the methylation levels of p21 and PPARγ (Abdalla et al, 2018;Qimuge et al, 2019). Zhang et al (2014) found that MBD4 inhibited porcine preadipocytes differentiation by changing the DNA methylation levels of adipogenic genes.…”

A Novel DNA Methyltransferase Dnmt3a3 Splice Variant Represses Preadipocyte Proliferation and Differentiation

“…Chicken primary preadipocytes were isolated from the abdominal fat tissue of 25-day-old yellow-feathered chicks (Kwangfeng) under sterile conditions as previously reported (Abdalla et al, 2018b). Briefly, the chicks (n = 3) were humanely slaughtered and the abdominal fat tissues (about 3 g) were rapidly excised, placed in a 6-cm petri dish containing 4 ml DMEM/F12 medium (Invitrogen, Carlsbad, United States), and minced into sections of approximately 1 mm 2 using scissors.…”

“…The novel transcript detection of Dnmt3a was performed as previously described (Abdalla et al, 2018b). Briefly, PCR amplification and Sanger sequencing methods were used to detect novel splice variants of Dnmt3a.…”

“…Dnmt3a2 is predominant in embryonic stem cells and is mainly localized to euchromatin, whereas Dnmt3a is primarily concentrated in the heterochromatin (Chen et al, 2002). Previously, we identified two transcripts of Dnmt3a, namely Dnmt3a and Dnmt3a1, in chicken (Abdalla et al, 2018b). The chicken Dnmt3a1 transcript was highly expressed in adipose tissue/ cell (Abdalla et al, 2018b) and its overexpression inhibits adipogenesis.…”

“…Previously, we identified two transcripts of Dnmt3a, namely Dnmt3a and Dnmt3a1, in chicken (Abdalla et al, 2018b). The chicken Dnmt3a1 transcript was highly expressed in adipose tissue/ cell (Abdalla et al, 2018b) and its overexpression inhibits adipogenesis. However, the biological roles of many spliced variants of a gene are not fully understood.…”