Reciprocal abrogation of PKM isoforms: contradictory outcomes and differing impact of splicing signal on CRISPR/Cas9 mediates gene editing in keratinocytes

• DOI: 10.1111/febs.16625
• Publication Date: 2022-09-09T21:19:14Z
• Authors: Simon P. Nold; Khrystyna Sych; Gergely Imre; Dominik C. Fuhrmann; Josef Pfeilschifter; Rajkumar Vutukuri; Frank Schnutgen; Ilka Wittig; Jana Meisterknecht; Stefan Frank; Itamar Goren
• Keywords: PKM2; HaCaT

The healing of wounded skin is a highly organized process involving massive cell in- and outflux, proliferation tissue remodelling. It well accepted that metabolic constraints such as diabetes mellitus, overweight or anorexia impairs wound healing. Indeed, inflammation involves boost overall changes. As converges inflammatory processes are also common to transformation, we investigate the functional role pro-neoplastic factor pyruvate kinase (PK) M2 its active splice variant PKM1 in keratinocytes. Particularly, challenge impact reciprocal ablation two expression. Here, CRISPR/Cas9 genome editing PKM gene HaCaT reveals an unexpected mutational bias at 3'SS exon 9, whereas no preference for any particular kind mutation 10 3' splice, despite close vicinity (400 nucleotides apart) sequence similarity between sites. Furthermore, opposed transient silencing PKM2, exclusion splicing PKM2 via mutually increases mRNA protein expression compensates absence elimination reduces impedes proliferation, thus unveiling essential growth balance