Abstract. Ubiquitylation of lysine residues in proteins serves a pivotal role in the efficient removal of misfolded or unused proteins and in the control of various regulatory pathways by monitoring protein activity that may lead to protein

Ubiquitylation of lysine residues in proteins serves a pivotal role in the efficient removal of misfolded or unused proteins and in the control of various regulatory pathways by monitoring protein activity that may lead to protein degradation. The loss of ubiquitylated lysines may affect the ubiquitin‐mediated regulatory network and result in the emergence of novel phenotypes. The present study analyzed mouse ubiquitylation data and orthologous proteins from 62 mammals to identify 193 conserved ubiquitylation sites from 169 proteins that were lost in the Euarchonta lineage leading to humans. A total of 8 proteins, including betaine homocysteine S‐methyltransferase, clin and CBS domain divalent metal cation transport mediator 3, ribosome‐binding protein 1 and solute carrier family 37 member 4, lost 1 conserved lysine residue, which was ubiquitylated in the mouse ortholog, following the human‐chimpanzee divergence. A total of 17 of the lost ubiquitylated lysines are also known to be modified by acetylation and/or succinylation in mice. In 8 cases, a novel lysine evolved at positions flanking the lost conserved lysine residues, potentially as a method of compensation. We hypothesize that the loss of ubiquitylation sites during evolution may lead to the development of advantageous phenotypes, which are then fixed by selection. The ancestral ubiquitylation sites identified in the present study may be a useful resource for investigating the association between loss of ubiquitylation sites and the emergence of novel phenotypes during evolution towards modern humans. Introduction Ubiquitylation, in which the highly conserved 76‐residue polypeptide ubiquitin is covalently attached to a lysine residue of substrate proteins, mediates the targeted destruction of ubiquitylated proteins by the ubiquitin‐proteasome system (1‐4). The ubiquitin‐mediated protein degradation pathway serves a crucial role in the efficient and specific removal of misfolded proteins and certain key regulatory proteins (5,6). Ubiquitin and other ubiquitin‐like proteins, including autophagy‐related protein 8, Ubiquitin‐like protein ISG15, NEDD8 conjugating enzyme UBE2F and SUMO‐conjugating enzyme UBC9, may all be targeted to the same lysine residue, and in this way they are involved in regulating a number of cellular processes including transcription, the cell cycle and signal transduction pathways (1,3,7,8). Modified lysine residues, covalently attached not only to ubiquitin but also to other ubiquitin‐like proteins, may be identified by high‐throughput techniques (9,10). In the present study, the term ‘ubiquitylation’ is used to refer to ubiquitin and other ubiquitin‐like protein modifications. The lysine residues that are targeted for ubiquitylation may also be modified by acetylation, and so cross‐talk between ubiquitylation and acetylation may control the stability of target proteins (11). It is also known that the cross‐talk between ubiquitylation and phosphorylation may modulate various regulatory networks (12). For example, ubiquitylation may positively or negatively regulate various protein kinases, either with or without their subsequent degradation (12‐14). A large number of genetic changes have accumulated during human evolution and are responsible for the acquisition of different human phenotypes (15,16). These genetic modifications include the de novo generation of genes (17), emergence of novel transcript isoforms (18) and loss of gene function (19‐21). The gain of novel phosphorylation sites in proteins may result in the reorganization of the regulatory circuits (22,23). The gain of lysine residues that are subject to ubiquitylation during human evolution may alter the timing of protein degradation, subcellular localization and the activity of the target proteins (24). Similarly, the gain or loss of N‐glycosylation modification sites in conserved proteins may have an effect on protein folding and trafficking, and on adhesion and cell signaling (25,26). In the present study, we hypothesized that the loss of conserved ubiquitylation sites in highly conserved proteins during evolution may modify the ubiquitin‐mediated regulatory network, which Loss of conserved ubiquitylation sites in conserved proteins during human evolution DONGBIN PARK, CHUL JUN GOH, HYEIN KIM, JI SEOK LEE and YOONSOO HAHN Department of Life Science, Chung‐Ang University, Seoul 06974, Republic of Korea Received January 30, 2018; Accepted July 6, 2018 DOI: 10.3892/ijmm.2018. 3772 Correspondence to: Professor Yoonsoo Hahn, Department of Life Science, Chung‐Ang University, 84 Heukseok‐ro, Dongjak‐gu, Seoul 06974, Republic of Korea E‐mail: hahny@cau.ac. kr Abbreviations: aa, amino acid; ER, endoplasmic reticulum; mUbiSiDa, mammalian ubiquitination site database; PTM, post‐translational modification; SNP, single nucleotide polymorphism; UPR, unfolded