Analysis of EHMT2 interactor isolation success using the minimal binding region

Abstract

Euchromatin Histone Lysine N-methyltransferase 2 (EHMT2) is a protein lysine methyltransferase (PKMT) that is known to have both histone and non-histone substrates and interactors (Tachibana et al., 2001; Rathert et al., 2008) and that has been found to be upregulated in various types of cancer (Casciello et al., 2015), with colorectal cancer (CRC) being one of them (Qin et al., 2018). EHMT2 has been observed to promote CRC progression in some contexts (Zhang et al., 2015; Zhang et al., 2018; Bergin et al., 2021) and to suppress it in others (Ichikawa et al., 2022), making further study into the enzyme’s role in CRC an important contributor to bettering CRC treatment and management. However, the full protein’s recombinant production has proved to be problematic. This study aims to determine whether or not EHMT2’s catalytic region can be used as a representative of the full protein in further studies by isolating its interactors using the yeast two hybrid assay, obtaining an indication of the interactions’ strength using the CPRG β-galactosidase assay, identifying the interactors via Sanger sequencing and comparing the isolated interactors with EHMT2’s known interactors. 7 potential interactors were identified via Sanger sequencing with the most promising ones being Monoamine Oxidase B, which is also highly expressed in CRC and associated with worse prognoses for patients (Yang et al., 2020), and Propionyl Coenzyme A Carboxylase Beta (PCCB), a component of PCC which is an enzyme that has been found to increase pro-metastatic markers in breast and lung cancer cells when overexpressed (Gomes et al., 2022). Further work is required to validate the specificity of these interactions.