bims-micpro 2021-10-31 papers

J Biomol Struct Dyn. 2021 Oct 27. 1-9

Microproteins: a 3D protein structure prediction analysis.

Kishan Thambu, Victoria Glomb, Rolando Hernandez Trapero, Julio C Facelli.

Microproteins are a novel and expanding group of small proteins encoded by less than 100-150 codons that are translated from small open reading frames (smORFs). It has been shown that smORFs and their corresponding microproteins make up a sizable fraction of the genome and proteome, but very little information on microproteins' structural features exists in the literature. In this paper, we present the results of analyzing the predicted structures of 44 microproteins. The results show that this set of microproteins have a different amino acid composition profiles, similar structural characteristics and fewer small-molecule ligand binding sites than regular proteins.Communicated by Ramaswamy H. Sarma.

Keywords: Gene ontology; ligand binding sites; microproteins; protein structure; protein structure prediction

DOI: https://doi.org/10.1080/07391102.2021.1993343

Bio Protoc. 2021 Sep 20. 11(18): e4167

Comprehensive Identification of Translatable Circular RNAs Using Polysome Profiling.

Yanwen Ye, Zefeng Wang, Yun Yang.

Circular RNAs (circRNAs), a special type of RNAs without 5'- and 3'-ends, are widely present in eukaryotes and known to function as noncoding RNAs to regulate gene expression, including as miRNA sponges. Recent studies showed that many exonic circRNAs, generated by back-splicing of pre-mRNAs, can be translated in a cap-independent fashion through IRESs or m6A RNA methylation. However, the scope of the translatable circRNAs and the biological function of their translation products are still unclear in different cells and tissues. Ribosome footprinting and proteomic analysis were usually used to globally identify translatable circRNAs. However, both methods have low sensitivity due to the low efficiency in the discovery of circRNA specific reads or peptides (i.e., the back-splicing junctions are difficult to recover by the short reads of ribosome footprinting and the limitation of proteomic analysis). Here, we described an alternative method to identify translatable circRNAs using polysome profiling and circRNA-seq. Generally, polysome-associated RNAs were separated with sucrose gradients. Then polysome-bound circRNAs were enriched by an RNase R treatment and identified through paired-end deep sequencing. Thus, this method is more sensitive than ribosome footprint and proteomic analyses for the identification of translatable circRNAs.

Keywords: Polysome profiling; RNA-seq; RNase R; back-splicing junction; circRNA translation

DOI: https://doi.org/10.21769/BioProtoc.4167

Methods Mol Biol. 2022 ;2404 331-351

High-Throughput Quantitation of Yeast uORF Regulatory Impacts Using FACS-uORF.

Gemma E May, C Joel McManus.

Eukaryotic upstream Open Reading Frames (uORFs) are short translated regions found in many transcript leaders (Barbosa et al. PLoS Genet 9:e1003529, 2013; Zhang et al. Trends Biochem Sci 44:782-794, 2019). Modern transcript annotations and ribosome profiling studies have found thousands of AUG-initiated uORFs, and many more uORFs initiated by near-cognate codons (CUG, GUG, UUG, etc.). Their translation generally decreases the expression of the main encoded protein by preventing ribosomes from reaching the main ORF of each gene, and by inducing nonsense mediated decay (NMD) through premature termination. Under many cellular stresses, uORF containing transcripts are de-repressed due to decreased translation initiation (Young et al. J Biol Chem 291:16927-16935, 2016). Traditional experimental evaluation of uORFs involves comparing expression from matched uORF-containing and start-codon mutated transcript leader reporter plasmids. This tedious process has precluded analysis of large numbers of uORFs. We recently used FACS-uORF to simultaneously assay thousands of yeast uORFs in order to evaluate the impact of codon usage on their functions (Lin et al. Nucleic Acids Res 2:1-10, 2019). Here, we provide a step-by-step protocol for this assay.

Keywords: Massively parallel reporter assay; mRNA translation; uORFs

DOI: https://doi.org/10.1007/978-1-0716-1851-6_18

OMICS. 2021 Oct 29.

The Normal Human Adult Hypothalamus Proteomic Landscape: Rise of Neuroproteomics in Biological Psychiatry and Systems Biology.

Oishi Chatterjee, Lathika Gopalakrishnan, Praseeda Mol, Jayshree Advani, Bipin Nair, Susarla Krishna Shankar, Anita Mahadevan, Thottethodi Subrahmanya Keshava Prasad.

The human hypothalamus is central to the regulation of neuroendocrine and neurovegetative systems, as well as modulation of chronobiology and behavioral aspects in human health and disease. Surprisingly, a deep proteomic analysis of the normal human hypothalamic proteome has been missing for such an important organ so far. In this study, we delineated the human hypothalamus proteome using a high-resolution mass spectrometry approach which resulted in the identification of 5349 proteins, while a multiple post-translational modification (PTM) search identified 191 additional proteins, which were missed in the first search. A proteogenomic analysis resulted in the discovery of multiple novel protein-coding regions as we identified proteins from noncoding regions (pseudogenes) and proteins translated from short open reading frames that can be missed using the traditional pipeline of prediction of protein-coding genes as a part of genome annotation. We also identified several PTMs of hypothalamic proteins that may be required for normal hypothalamic functions. Moreover, we observed an enrichment of proteins pertaining to autophagy and adult neurogenesis in the proteome data. We believe that the hypothalamic proteome reported herein would help to decipher the molecular basis for the diverse range of physiological functions attributed to it, as well as its role in neurological and psychiatric diseases. Extensive proteomic profiling of the hypothalamic nuclei would further elaborate on the role and functional characterization of several hypothalamus-specific proteins and pathways to inform future research and clinical discoveries in biological psychiatry, neurology, and system biology.

Keywords: biological psychiatry; brain research; hypothalamus; neuropeptides; post-translational modifications; proteomics

DOI: https://doi.org/10.1089/omi.2021.0158