Coactivator Associated Arginine Methyltransferase 1 (CARM1) as a Contributor to Motoneuron Biology

Abstract

Coactivator Associated Arginine Methyltransferase 1 (CARM1) is a ubiquitously expressed protein arginine methyltransferase with essential roles in normal cell function. As such, CARM1 is often a part of the investigative rationale, where its quantification through Western Blot (WB) is commonly assessed. Here, CARM1 is shown to form aggregates that prevent its migration in SDS-PAGE and introduces error in its quantification by WB. Heat denaturation, DTT and CARM1’s own concentration in samples are found to contribute to its aggregation. An adapted sample buffer with higher SDS concentrations and without DTT prevents aggregation and permits accurate quantification. These findings reframe CARM1’s biochemical properties as an essential component of its study in an experimental and potentially biological setting. In motoneurons (MNs), CARM1 was previously found to methylate HuD, a major neuronal RNA binding protein, thereby altering its affinity with the p21 mRNA. Here, the notion that other mRNAs are similarly impacted was explored, and Schip1/Iqschfp, Smarca2, Stc2, Ptger4 and Phox2B mRNAs were found to be dependent on CARM1 levels for their interaction with HuD. These interactions were not previously characterized, making them novel elements in both HuD’s and CARM1’s biology. Moreover, the protein levels of Ptger4 and Smarca2 were regulated by CARM1’s enzymatic activity expanding and attesting to its functional relevance in MNs. Akin to other work, CARM1’s interactome identifies Alix as being by far its most prominent interactor. Also, ribosomal proteins are overrepresented in CARM1’s interactome suggesting that translation remains a major facet of its function in MNs. In line with this assessment, this work identifies RPS7 as a novel substrate of CARM1. This study solidifies CARM1 as a contributor to neuronal biology through its regulation of HuD mRNA interactions, its regulation of Ptger4 and Smarca2 protein levels and contribution to the MN methylome.