Thacker G, Mishra M, Sharma A, Singh AK, Sanyal S, Trivedi AK

Transcription factor CCAAT/Enhancer binding protein alpha (C/EBP?) is a key regulator of myeloid differentiation, granulopoiesis in particular. Although CEBPA mutations are found in more than 10% in AML, functional inhibition of C/EBP? protein is also widely observed in AML. Here, we sought to examine if SKP2, an aberrantly enhanced E3 ...

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ubiquitin ligase in primary AMLs inhibits C/EBP? stability to induce differentiation block.Here we employed cell based assays such as transfections, immunoblotting, co-immunoprecipitation, luciferase and gel shift assays along with differentiation assays to investigate SKP2 regulated C/EBP? protein stability in acute myeloid leukemia.Here we discovered that oncogenic E3 ubiquitin ligase SCFskp2 ubiquitinates and destabilizes C/EBP? in a proteasome-dependent manner. Our data demonstrates that SKP2 physically interacts with C-terminal of C/EBP? and promotes its K48-linked ubiquitination-mediated degradation leading to its reduced transactivation potential, DNA binding ability and cellular functions. We further show that while overexpression of SKP2 inhibits both ectopic as well as endogenous C/EBP? in heterologous (HEK293T) as well as myeloid leukemia cells respectively, SKP2 depletion restores endogenous C/EBP? leading to reduced colony formation and enhanced myeloid differentiation of myeloid leukemia cells. Using Estradiol-inducible K562-C/EBP?-ER cells as yet another model of granulocytic differentiation, we further confirmed that SKP2 overexpression indeed inhibits granulocytic differentiation by mitigating C/EBP? stability.Our findings identify SKP2 as a potential negative regulator of C/EBP? stability and function in AML which suggests that SKP2 can be potentially targeted in AML to restore C/EBP? and overcome differentiation block.

Date: Sep. 15, 2020

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