Somatic mutations in p53, which inactivate the tumour-suppressor function of p53 and often confer oncogenic gain-of-function properties, are very common in cancer 1 , 2 . Here we studied the effects of hotspot gain-of-function mutations in Trp53 (the gene that encodes p53 in mice) in mouse models of WNT-driven intestinal cancer caused by Csnk1a1 deletion 3 , 4 or Apc Min mutation 5 . Cancer in these models is known to be facilitated by loss of p53 3 , 6 . We found that mutant versions of p53 had contrasting effects in different segments of the gut: in the distal gut, mutant p53 had the expected oncogenic effect; however, in the proximal gut and in tumour organoids it had a pronounced tumour-suppressive effect. In the tumour-suppressive mode, mutant p53 eliminated dysplasia and tumorigenesis in Csnk1a1- deficient and Apc Min /+ mice, and promoted normal growth and differentiation of tumour organoids derived from these mice. In these settings, mutant p53 was more effective than wild-type p53 at inhibiting tumour formation. Mechanistically, the tumour-suppressive effects of mutant p53 were driven by disruption of the WNT pathway, through preventing the binding of TCF4 to chromatin. Notably, this tumour-suppressive effect was completely abolished by the gut microbiome. Moreover, a single metabolite derived from the gut microbiota–gallic acid–could reproduce the entire effect of the microbiome. Supplementing gut-sterilized p53-mutant mice and p53-mutant organoids with gallic acid reinstated the TCF4–chromatin interaction and the hyperactivation of WNT, thus conferring a malignant phenotype to the organoids and throughout the gut. Our study demonstrates the substantial plasticity of a cancer mutation and highlights the role of the microenvironment in determining its functional outcome.

Using oxygen and NADPH, the redox enzymes cytochrome P450 (CYP) and its reductase (CPR) work in tandem to carry out the phase I metabolism of a vast majority of drugs and xenobiotics. As per the erstwhile understanding of the catalytic cycle, binding of the substrate to CYP's heme distal pocket allows CPR to pump electrons through a CPR-CYP complex. In turn, this trigger (a thermodynamic push of electrons) leads to the activation of oxygen at CYP's heme-center, to give Compound I, a two-electron deficient enzyme reactive intermediate. The formation of diffusible radicals and reactive oxygen species (DROS, hitherto considered an undesired facet of the system) was attributed to the heme-center. Recently, we had challenged these perceptions and proposed the murburn (“mured burning” or “mild unrestricted burning”) concept to explain heme enzymes' catalytic mechanism, electron-transfer phenomena and the regulation of redox equivalents' consumption. Murburn concept incorporates a one-electron paradigm, advocating obligatory roles for DROS. The new understanding does not call for high-affinity substrate-binding at the heme distal pocket of the CYP (the first and the most crucial step of the erstwhile paradigm) or CYP-CPR protein-protein complexations (the operational backbone of the erstwhile cycle). Herein, the dynamics of reduced nicotinamide nucleotides' consumption, peroxide formation and depletion, product(s) formation, etc. was investigated with various controls, by altering reaction variables, environments and through the incorporation of diverse molecular probes. In several CYP systems, control reactions lacking the specific substrate showed comparable or higher peroxide in milieu, thereby discrediting the foundations of the erstwhile hypothesis. The profiles obtained by altering CYP:CPR ratios and the profound inhibitions observed upon the incorporation of catalytic amounts of horseradish peroxidase confirm the obligatory roles of DROS in milieu, ratifying murburn as the operative concept. The mechanism of uncoupling (peroxide/water formation) was found to be dependent on multiple one and two electron equilibriums amongst the reaction components. The investigation explains the evolutionary implications of xenobiotic metabolism, confirms the obligatory role of diffusible reactive species in routine redox metabolism within liver microsomes and establishes that a redox enzyme like CYP enhances reaction rates (achieves catalysis) via a novel (hitherto unknown) modality.

No abstract

CKIα ablation induces p53 activation, and CKIα degradation underlies the therapeutic effect of lenalidomide in a pre-leukemia syndrome. Here we describe the development of CKIα inhibitors, which co-target the transcriptional kinases CDK7 and CDK9, thereby augmenting CKIα-induced p53 activation and its anti-leukemic activity. Oncogene-driving super-enhancers (SEs) are highly sensitive to CDK7/9 inhibition. We identified multiple newly gained SEs in primary mouse acute myeloid leukemia (AML) cells and demonstrate that the inhibitors abolish many SEs and preferentially suppress the transcription elongation of SE-driven oncogenes. We show that blocking CKIα together with CDK7 and/or CDK9 synergistically stabilize p53, deprive leukemia cells of survival and proliferation-maintaining SE-driven oncogenes, and induce apoptosis. Leukemia progenitors are selectively eliminated by the inhibitors, explaining their therapeutic efficacy with preserved hematopoiesis and leukemia cure potential; they eradicate leukemia in MLL-AF9 and Tet2;Flt3 AML mouse models and in several patient-derived AML xenograft models, supporting their potential efficacy in curing human leukemia.

Reductase reduces cytochrome c via relays of highly mobile diffusible agents; not by direct binding and inter-protein long-distance electron tunnelling.