Search

As the most widespread mRNAs modification, N6-methyladenosine (mHerein, we discovered that in CRC, downregulated ALKBH5 was closely related to poor prognosis of CRC patients. Functionally, our results demonstrated that knockdown of ALKBH5 enhanced the proliferation, migration and invasion of LOVO and RKO in vitro, while overexpression of ALKBH5 inhibited the functions of these cells. The results also demonstrated that knockdown of ALKBH5 promoted subcutaneous tumorigenesis of LOVO in vivo, while overexpression of ALKBH5 suppressed this ability. Mechanistically, results from joint analyses of MeRIP-seq and RNA-seq indicated that PHF20 mRNA was a key molecule that was regulated by ALKBH5-mediated mALKBH5 suppresses CRC progression by decreasing PHF20 mRNA methylation. ALKBH5-mediated m

The intervertebral disc (IVD) degeneration is the leading cause of low back pain, which accounts for a main cause of disability. N6-methyladenosine (m6A) is the most abundant internal modification in eukaryotic messenger RNAs and is involved in various diseases and cellular processes by modulating mRNA fate. However, the critical role of m6A regulation in IVD degeneration remains unclear. Nucleus pulposus cell (NPC) senescence is critical for the progression of IVD degeneration. Here, we uncovered the role and explored the regulatory mechanism of m6A in NPC senescence during IVD degeneration.Identification of NPC senescence during IVD degeneration was based on the analysis of tissue samples and the cellular model. ALKBH5 upregulation inducing cellular senescence was confirmed by functional experiments in vivo and in vitro. ChIP-qPCR and DNA-Pulldown were used to reveal increased ALKBH5 was regulated by KDM4A-mediated H3K9me3. Furthermore, Me-RIP-seq was performed to identify m6A hypomethylation of DNMT3B transcripts in senescent NPCs. Stability analysis showed that DNMT3B expression was enhanced for less YTHDF2 recognition and increased DNMT3B promoted NPC senescence and IVD degeneration via E4F1 methylation by in vivo and in vitro analyses.Expression of ALKBH5 is enhanced during IVD degeneration and NPC senescence, due to decreased KDM4A-mediated H3K9me3 modification. Functionally, ALKBH5 causes NPC senescence by demethylating DNMT3B transcripts and in turn promoting its expression via less YTHDF2 recognition and following degradation due to transcript hypomethylation in vitro and in vivo. Increased DNMT3B promotes the development of IVD degeneration and NPC senescence, mechanistically by methylating CpG islands of E4F1 at the promoter region and thus restraining its transcription and expression.Collectively, our findings reveal an epigenetic interplay mechanism in NPC senescence and IVD degeneration, presenting a critical pro-senescence role of ALKBH5 and m6A hypomethylation, highlighting the therapeutic potential of targeting the m6A/DNMT3B/E4F1 axis for treating IVD degeneration.

Background Post‐ischemic angiogenesis is critical for blood flow recovery and ischemic tissue repair. N6‐methyladenosine (m6A) plays essential roles in numerous biological processes. However, the impact and connected mechanism of m6A on post‐ischemic angiogenesis are not fully understood. Methods AlkB homolog 5 (ALKBH5) was screened out among several methyltransferases and demethylases involved in dynamic m6A regulation. Cardiac microvascular endothelial cells (CMECs) angiogenesis and WNT family member 5A (WNT5A) stability were analyzed upon ALKBH5 overexpression with adenovirus or knockdown with small interfering RNAs in vitro. The blood flow recovery, capillary, and small artery densities were evaluated in adeno‐associated virus (AAV)‐ALKBH5 overexpression or ALKBH5 knockout (KO) mice in a hind‐limb ischemia model. The same experiments were conducted to explore the translational value of transient silencing of ALKBH5 with adenovirus. Results ALKBH5 was significantly upregulated in hypoxic CMECs and led to a global decrease of m6A level. ALKBH5 overexpression further reduced m6A level in normoxic and hypoxic CMECs, impaired proliferation, migration, and tube formation only in hypoxic CMECs. Conversely, ALKBH5 knockdown preserved m6A levels and promoted angiogenic phenotypes in hypoxic but not in normoxic CMECs. Mechanistically, ALKBH5 regulated WNT5A expression through post‐transcriptional mRNA modulation in an m6A‐dependent manner, which decreased its stability and subsequently impeded angiogenesis in hypoxic CMECs. Furthermore, ALKBH5 overexpression hindered blood flow recovery and reduced CD31 and alpha‐smooth muscle actin expression in hind‐limb ischemia mice. As expected, ALKBH5‐KO mice exhibited improved blood flow recovery, increased capillary, and small artery densities after hind‐limb ischemia, and similar beneficial effects were observed in mice with transient adenoviral ALKBH5 gene silencing. Conclusion We demonstrate that ALKBH5 is a negative regulator of post‐ischemic angiogenesis via post‐transcriptional modulation and destabilization of WNT5A mRNA in an m6A‐dependent manner. Targeting ALKBH5 may be a potential therapeutic option for ischemic diseases, including peripheral artery disease.