3.3 | Ubiquitin-like microproteins
Several groups recently reported the discovery of ubiquitin-like microproteins. In one example, the ubiquitin pseudogene UBBP4 was reported to be translated. Interestingly, UBBP4 encodes three ubiquitin variants within two independent open reading frames, and mass spectrometric evidence uniquely identifying all three have been previously obtained. The UBBP4 ubiquitin-like proteins exhibit high sequence similarity to canonical ubiquitin, with 1 (variant Ubbp4A1), 4 (Ubbp4B1 or UbKEKS), or 8 amino acid substitutions (Ubbp4A2). Ubbp4A2 and UbKEKS retain a functional C-terminal diglycine motif and can be covalently conjugated to high molecular weight cellular proteins, while UbbpA1 was predominantly observed as a monomer. Despite being ~700-fold less abundant than canonical ubiquitin, UbKEKS modifies a specific subset of cellular proteins including lamins, and, rather than promoting proteasomal degradation, may be important for regulating target protein localization and/or function.
In 2020 the TINCR RNA, which was previously classified as noncoding, was shown to encode an 87-amino acid microprotein with 85% sequence homology to ubiquitin. The microprotein translated fromTINCR RNA, termed pTINCR or TUBL, was predicted to adopt a ubiquitin-like fold (Figure 4D). This prediction was confirmed in a recent crystal structure of pTINCR, which revealed an overall ubiquitin-like fold with a positively charged N-terminal domain hypothesized to enable interaction with other biomolecules (Figure 4E). Due to the lack of a C-terminal diglycine motif, pTINCR is a type II ubiquitin-like protein that associates with ubiquitin-binding proteins rather than being covalently attached to proteins. pTINCR is expressed in skin, and mice lacking pTINCR exhibit a mild delay in wound healing. Importantly, two reports have identified pTINCR as a tumor suppressor in cutaneous squamous cell carcinoma and other epithelial cancers. pTINCR is upregulated after DNA damage-induced p53 activation, and it is frequently lost or mutated in squamous cell carcinoma. It normally promotes differentiation of keratinocytes and other epithelial cell types via its interaction with SUMOylated Cdc42. Consequently, mouse embryonic stem cell-derived teratomas overexpressing pTINCR exhibit decreased growth and increased keratin deposition consistent with involvement in differentiation of skin cells. Along the same lines, pTINCR overexpression inhibits the proliferation of squamous cell carcinoma cells in culture and in xenografts. Additionally, mice heterozygous for Xpc that lack pTINCR are DNA damage repair-deficient and exhibit increased formation of invasive skin papillomas and squamous cell carcinomas relative to Xpcheterozygous/pTINCR wild-type mice upon UV exposure. Overall, pTINCR is a type II ubiquitin-like microprotein that is required for keratinocyte differentiation and acts as a tumor suppressor in squamous cell carcinoma.