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Open Access | 
10.1172/JCI179560
VHL governs m6A modification and PIK3R3 mRNA stability in clear cell renal cell carcinomas
Hyemin Lee,1 Li Zhuang,1 and Boyi Gan1,2
1Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
2The University of Texas MD Anderson UTHealth Graduate School of Biomedical Sciences, Houston, Texas, USA.
Address correspondence to: Boyi Gan, Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, 6565 MD Anderson Blvd, Houston, TX, 77030, USA. Phone: 713.792.8653; Email: bgan@mdanderson.org.
Find articles by Lee, H. in: JCI | PubMed | Google Scholar
1Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
2The University of Texas MD Anderson UTHealth Graduate School of Biomedical Sciences, Houston, Texas, USA.
Address correspondence to: Boyi Gan, Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, 6565 MD Anderson Blvd, Houston, TX, 77030, USA. Phone: 713.792.8653; Email: bgan@mdanderson.org.
Find articles by Zhuang, L. in: JCI | PubMed | Google Scholar
1Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA.
2The University of Texas MD Anderson UTHealth Graduate School of Biomedical Sciences, Houston, Texas, USA.
Address correspondence to: Boyi Gan, Department of Experimental Radiation Oncology, The University of Texas MD Anderson Cancer Center, 6565 MD Anderson Blvd, Houston, TX, 77030, USA. Phone: 713.792.8653; Email: bgan@mdanderson.org.
Find articles by Gan, B. in: JCI | PubMed | Google Scholar
Published April 15, 2024 - More info
J Clin Invest. 2024;134(8):e179560. https://doi.org/10.1172/JCI179560.
© 2024 Lee et al. This work is licensed under the Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
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Abstract
N6-Methyladenosine (m6A) is the most abundant posttranscriptional modification, and its contribution to cancer evolution has recently been appreciated. Renal cancer is the most common adult genitourinary cancer, approximately 85% of which is accounted for by the clear cell renal cell carcinoma (ccRCC) subtype characterized by VHL loss. However, it is unclear whether VHL loss in ccRCC affects m6A patterns. In this study, we demonstrate that VHL binds and promotes METTL3/METTL14 complex formation while VHL depletion suppresses m6A modification, which is distinctive from its canonical E3 ligase role. m6A RNA immunoprecipitation sequencing (RIP-Seq) coupled with RNA-Seq allows us to identify a selection of genes whose expression may be regulated by VHL-m6A signaling. Specifically, PIK3R3 is identified to be a critical gene whose mRNA stability is regulated by VHL in a m6A-dependent but HIF-independent manner. Functionally, PIK3R3 depletion promotes renal cancer cell growth and orthotopic tumor growth while its overexpression leads to decreased tumorigenesis. Mechanistically, the VHL-m6A–regulated PIK3R3 suppresses tumor growth by restraining PI3K/AKT activity. Taken together, we propose a mechanism by which VHL regulates m6A through modulation of METTL3/METTL14 complex formation, thereby promoting PIK3R3 mRNA stability and protein levels that are critical for regulating ccRCC tumorigenesis.
Authors
Cheng Zhang, Miaomiao Yu, Austin J. Hepperla, Zhao Zhang, Rishi Raj, Hua Zhong, Jin Zhou, Lianxin Hu, Jun Fang, Hongyi Liu, Qian Liang, Liwei Jia, Chengheng Liao, Sichuan Xi, Jeremy M. Simon, Kexin Xu, Zhijie Liu, Yunsun Nam, Payal Kapur, Qing Zhang
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m6A RNA modification in VHL-deficient ccRCCs
N6-methyladenosine (m6A) represents one of the most prevalent RNA chemical modifications in mRNA that substantially impacts RNA fate and expression by modulating cellular processes such as mRNA splicing, stability, nuclear export, and translation (7). In recent years, numerous studies have shed light on the crucial role of m6A in governing tumor proliferation, invasion, and metastasis, including in ccRCCs (8). Several investigations suggest that METTL14, a critical component of the m6A methyltransferase writer complex alongside METTL3 and WTAP, functions as a tumor suppressor in ccRCCs and highlight its role in attenuating the proliferation and migration capabilities of renal cancer cells (9, 10). However, the extent and specific mechanisms through which the m6A regulatory machinery is influenced by key oncogenic events in ccRCCs have remained elusive.
In this issue of the JCI, Zhang and colleagues have uncovered a role of VHL in regulating m6A modification and RNA stability in ccRCCs (11). Mechanistically, VHL interacted with proteins of the m6A enzymatic complex and modulated the interaction between METTL3 and METTL14, thereby regulating m6A levels in RCC cells (Figure 1). Depletion of VHL resulted in decreased global m6A levels in VHL-proficient renal cancer cells, with this regulation being mediated by VHL’s E3 ligase domain. Interestingly, VHL’s effect on m6A levels seems to operate independently of its canonical function in regulating HIFs. For example, the authors demonstrated that the overexpression of a HIF2α mutant that is resistant to VHL-mediated degradation in VHL-expressing RCC cells — in which the WT HIF2α protein is constitutively degraded by VHL — did not reverse the effect of VHL on m6A levels. In addition, contrary to VHL restoration, the deletion of HIF2α in RCC cells — which exhibit elevated HIF2α protein levels due to VHL loss — did not impact m6A levels.
Figure 1VHL mediates m6A RNA modification in ccRCCs. VHL orchestrates the complex assembly of METTL3 and METTL14, leading to the incorporation of m6A into PIK3R3 mRNA. The m6A reader proteins, IGF2BP1 and IGF2BP2, play a role in stabilizing PIK3R3 mRNA. The stabilized PIK3R3 mRNA leads to increased levels of PIK3R3 protein, which in turn exert a negative regulatory effect on the p85 protein, consequently inhibiting PI3K/AKT signaling and impeding ccRCC tumorigenesis.
Rigorous m6A RNA immunoprecipitation–sequencing (MeRIP-Seq) analyses revealed numerous differentially regulated m6A sites upon VHL depletion (11). In conjunction with RNA-Seq, Zhang et al. identified genes exhibiting differential m6A modification and expression changes upon VHL depletion. Notably, among these genes, the authors identified phosphoinositide-3-kinase regulatory subunit 3 (PIK3R3) as one of the implicated targets. Upon VHL depletion, the degradation rate of PIK3R3 mRNA accelerated the decay process of PIK3R3 mRNA. Conversely, overexpression of VHL decelerated its decay process. Consequently, VHL plays a critical role in maintaining the RNA stability of PIK3R3. Furthermore, IGF2BP1 and IGF2BP2 were identified as the primary m6A readers governing the regulation of PIK3R3 mRNA. These studies suggest a model in which VHL facilitates the assembly of the METTL3 and METTL14 complex and increases m6A occupancy on PIK3R3 mRNA, enhancing the stability of PIK3R3 mRNA and consequently its protein levels (Figure 1).
Zhang and colleagues subsequently identified PIK3R3 as a regulator of the ubiquitination process involving the PI3K regulatory protein p85. Additionally, they demonstrated that p85α played a positive role in AKT activation (11), contrary to its function as a tumor suppressor in some other cancer types (12). PIK3R3 mRNA stabilization by VHL through m6A modification resulted in elevated protein levels of PIK3R3, which then negatively regulated p85 and attenuated PI3K/AKT activation, subsequently impeding renal tumor growth. Conversely, VHL loss in ccRCC cells decreased PIK3R3 levels, resulting in AKT hyperactivation and enhanced renal tumor growth (Figure 1).
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ISSN: 0021-9738 (print), 1558-8238 (online)