Dual-stress-responsive-pathways prevent HCC initiation via p53 suppression of MTHFD1L-RNA m⁶A-elicited autonomous...

Hepatocellular carcinoma (HCC) originates from damaged hepatocytes in chronic liver injury. Though HCC development requires oncoproteins-mediated p53 inhibition, both genetic deletion and continous activation of p53 specifically in mouse hepatocytes are shown to promote liver tumorigenesis, suggesting that chronic liver injury firstly activates p53, and p53 activity needs to be dynamically regulated for preventing hepatocarcinogenesis. Yet, it remains unaddressed how liver injury stresses trigger signaling pathways to rapidly activate p53 prior to its inactivation in a physiological setting. Two pathways, ribosomal proteins (RPs)-MDM2 and 14-3-3-MDMX, are shown to activate p53 upon stresses, and mutations of MDM2 or MDMX that may disrupt these pathways have been found in human cancers including HCC. Using our unique double knock-in (DKI) mice that contain wild-type p53 while harbor defects in these two pathways, we unveiled that basal level of p53 in DKI mice is sufficient for maintaining liver function, however, carcinogen- or unhealthy diet-induced HCC initiation is accelerated in DKI mice. The two p53-pathways are also activated in human cirrhotic livers. We futher identified a one-carbon metabolism (1CM) enzyme methylenetetrahydrofolate dehydrogenase 1 like (MTHFD1L) as the p53-suppressed player in HCC initiation. We found that MTHFD1L is upregulated in human early HCC and correlated with p53 status, and promotes mouse HCC initiation by enhancing autonomous growth and immune evasion of HCC initiating cells (HCICs). With Nanopore RNA-m⁶A-sequencing, we unveiled that MTHFD1L fulfills these bi-functions by fueling methionine cycle that produces methyl groups to maintain the mRNA m⁶A of Snail (an HCC early tumorigenesis driver) and B2m (a light chain of MHC-I required for antigen presentation to CD8+ T) in HCICs. Our findings demonstrate that the two stress-triggered p53-pathways are crucial for preventing liver early tumorigenesis by reversing MTHFD1L upregulation-associated 1CM reprogramming and subsequent Snail and B2m mRNA-m⁶A-mediated cell autonomous growth and immune evasion.