Although long noncoding RNAs (lncRNAs) are by definition noncoding, they are known to encode disproportionately short molecules—mere polypeptides, not proteins. Whether these polypeptides matter very much is another question. They could represent nothing more than transcriptional noise, or they could be—as a new study suggests—functionally relevant.
In the new study, scientists based at Beth Israel Deaconess Medical Center (BIDMC) computationally predicted and experimentally confirmed the presence of “many” lncRNA-encoded polypeptides that were once “hidden.” More importantly, the scientists demonstrated that at least one of these polypeptides is functionally relevant.
This peptide, a 90-amino-acid-long molecule encoded by the lncRNA LINC00961, is called Small regulatory Polypeptide of Amino acid Response (SPAR). It appears to play an important role in modulating the activity of the mechanistic target of rapamycin complex 1 (mTORC1) protein complex, which is a critical sensor of nutrient availability within cells. The complex also regulates a variety of cellular processes, including translation, metabolism, cell growth, and proliferation; alterations in its function can lead to diseases such as cancer.
The new findings, which appeared December 26 in a paper (“mTORC1 and Muscle Regeneration Are Regulated by the LINC00961-Encoded SPAR Polypeptide”) that appeared in Nature, suggest that lncRNAs are functionally relevant. Specifically, they may help control cellular components in a tissue-specific manner.
“We show that the SPAR-encoding lncRNA is highly expressed in a subset of tissues and use CRISPR/Cas9 engineering to develop a SPAR-polypeptide-specific knockout mouse while maintaining expression of the host lncRNA,” wrote the paper’s authors. “We find that the SPAR-encoding lncRNA is downregulated in skeletal muscle upon acute injury, and using this in vivo model we establish that SPAR downregulation enables efficient activation of mTORC1 and promotes muscle regeneration.”
The BIDMC scientists, led by the Nature paper’s senior author, Pier Paolo Pandolfi, M.D., Ph.D., used computational analyses to predict potential polypeptides that could be encoded by known lncRNA molecules, and then they used mass spectrometry to determine if these putative polypeptides were actually expressed. After uncovering a number of novel polypeptides, they focused on SPAR. This polypeptide, they found, is expressed in a number of tissue types, including muscle.
Experiments conducted in mice demonstrated that through its effects on mTORC1, the SPAR polypeptide helps regulate the muscle's ability to regenerate and repair after injury. Specifically, expression of LINC00961 is blocked following muscle injury in mice, leading to reduced levels of SPAR and maximal mTORC1 activity to promote tissue regeneration.
“The experimental approach we used allowed us to eliminate expression of the SPAR polypeptide, while maintaining expression of the host lncRNA,” said lead author Akinobu Matsumoto, Ph.D., research fellow at the Cancer Center at BIDMC and lead author of the study. “We are able to ascribe this function to the coding function of the lncRNA rather than any noncoding function it may also have.” The findings suggest that therapeutic strategies that restrict expression of SPAR in injured muscle may promote a more rapid regeneration of tissue.
“Additionally, and more generally,” the paper’s authors noted, “the [new findings] emphasize that lncRNA-encoded polypeptides are more than just translational noise and can encode functionally relevant polypeptides with clinical and therapeutic relevance.” Basically, lncRNAs may have diverse roles and functions. Although they may not code for large proteins, lncRNAs may produce small polypeptides that can fine-tune the activity of critical cellular components.
The authors of the current study promise that they will, in a separate paper, provide a detailed description of the proteomics strategy they used to identify novel polypeptides encoded by putative lncRNAs. For now, however, they indicated that they have already expanded the repertoire of peptide-coding genes in the human genome that should be studied and annotated.
“Our team set about trying to understand to what extent lncRNA molecules might actually encode functional polypeptides and how important such peptides might be,” asserted Dr. Pandolfi. Arguably, the team achieved just that. Also, by establishing SPAR’s activation of mTORC1, the team provided a “paradigm by which lncRNAs encoding small polypeptides can modulate general biological pathways and processes to facilitate tissue-specific requirements, consistent with their restricted and highly regulated expression profile.”
“An ability to target modulators such as mTORC1 could be of great advantage from a therapeutic perspective, allowing for control of mTORC1 activity in cells or tissues that express such modulators while not affecting its activity and function in other tissue and cell types,” explained co-author John Clohessy, Ph.D., instructor in medicine at BIDMC and a senior member of Dr. Pandolfi's research team. Indeed, a key feature of many lncRNAs is that their expression is often highly tissue specific. Thus, targeting small polypeptides encoded by lncRNA molecules may provide the key to regulating common cellular components in a tissue-specific manner.
Because the mTORC1 complex is frequently deregulated in conditions such as cancer, the research team is now seeking to determine if SPAR can influence additional cellular functions of mTORC1 that might be involved in different diseases. “We are very excited about this discovery,” concluded Dr. Pandolfi. “It represents a new and startling mechanism by which important sensory pathways can be regulated within cells, and we believe it will have important implications for how we approach the design of therapies and treatments in the future.”