bims-micpro 2023-10-08 papers

Curr Opin Microbiol. 2023 Sep 28. pii: S1369-5274(23)00121-2. [Epub ahead of print]76 102384

The power of the small: the underestimated role of small proteins in bacterial and archaeal physiology.

Britta Jordan, Katrin Weidenbach, Ruth A Schmitz.

Small proteins encoded by small open-reading frames (sORFs) (≤70 aa) were overlooked for decades due to methodological reasons and are thus often missing in genome annotations. Novel detection methods such as ribosome profiling (Ribo-Seq) and mass spectrometry optimized for small proteins (peptidomics) have opened up a new field of interest and several catalogs of small proteins in bacteria and archaea have been recently reported. Many translated sORFs have been discovered in genomic locations previously thought to be noncoding, such as 5' or 3' untranslated regions or well-studied regulatory small RNAs (sRNAs). Even within longer ORFs, additional functional sORFs have been detected. Today, only a small proportion is characterized, but those small proteins indicate important and diverse functions in cellular physiology. Here, we summarize recently characterized small proteins involved in microbial metabolism.

DOI: https://doi.org/10.1016/j.mib.2023.102384

Microbiol Spectr. 2023 Oct 02. e0252823

Identification of proteoforms of short open reading frame-encoded peptides in Blautia producta under different cultivation conditions.

Jerome Genth, Kathrin Schäfer, Liam Cassidy, Simon Graspeuntner, Jan Rupp, Andreas Tholey.

Short open reading frames (sORF) have the potential to encode small proteins, sORF-encoded peptides (SEP), which are less than or equal to 100 amino acids in length. These SEP have been associated with various physiological processes and are known for their potential roles in modulating immune responses, signaling pathways, and cellular functions. Using bottom-up and top-down proteomics, we identified 45 novel and formerly reported SEP under different growth conditions and stress exposure in single cultures of Blautia producta that plays a central role in mediating colonization resistance and dampening inflammation in the human gut microbiome. Top-down analysis improved identification confidence and allowed the detection of a number of full-length and N- or C-terminal truncated proteoforms of the SEP and of proteoforms carrying post-translational modifications such as disulfides, which provides hints for a wide functional potential of these molecules. The identification of the SEP in single cultures of this bacterium shows that, in contrast to previous results, the biosynthesis of specific SEP is not restricted to direct bacterial interactions within the microbiome but is dependent on growth and environmental conditions during cultivation. IMPORTANCE The identification of short open reading frame-encoded peptides (SEP) and different proteoforms in single cultures of gut microbes offers new insights into a largely neglected part of the microbial proteome landscape. This is of particular importance as SEP provide various predicted functions, such as acting as antimicrobial peptides, maintaining cell homeostasis under stress conditions, or even contributing to the virulence pattern. They are, thus, taking a poorly understood role in structure and function of microbial networks in the human body. A better understanding of SEP in the context of human health requires a precise understanding of the abundance of SEP both in commensal microbes as well as pathogens. For the gut beneficial B. producta, we demonstrate the importance of specific environmental conditions for biosynthesis of SEP expanding previous findings about their role in microbial interactions.

Keywords: gut microbiome; microproteins; proteoforms; sORF-encoded peptides; short open reading frames; small open reading frames; top-down proteomics

DOI: https://doi.org/10.1128/spectrum.02528-23

Mol Plant. 2023 Sep 29. pii: S1674-2052(23)00291-5. [Epub ahead of print]

Plant and animal positive-sense single-stranded RNA viruses encode small proteins important for viral infection in their negative-sense strand.

Pan Gong, Qingtang Shen, Mingzhen Zhang, Rui Qiao, Jing Jiang, Lili Su, Siwen Zhao, Shuai Fu, Yu Ma, Linhao Ge, Yaqin Wang, Rosa Lozano-Durán, Aiming Wang, Fangfang Li, Xueping Zhou.

Positive-sense single-stranded RNA (+ssRNA) viruses, the most abundant viruses of eukaryotes in nature, require the synthesis of negative-sense RNA (-RNA) using their genomic (positive-sense) RNA (+RNA) as a template for replication. Based on the current evidence, viral proteins are translated via viral positive sense RNAs, whereas -RNA is considered to be a viral replication intermediate without coding capacity. Here, we report that plant and animal +ssRNA viruses contain small open reading frames (ORFs) in their -RNA (rORFs). Using turnip mosaic virus (TuMV) as a model for plant +ssRNA viruses, we demonstrate that small proteins encoded by rORFs display specific subcellular localizations, and confirm the presence of rORF2 in infected cells through mass spectrometry analysis. The protein encoded by TuMV rORF2 forms punctuate granules that localize in the perinuclear region and co-localize with viral replication complexes. The rORF2 protein can directly interact with the viral RNA-dependent RNA polymerase; mutation of rORF2 completely abolishes virus infection, and ectopic expression of rORF2 rescues the mutant virus. Furthermore, we show that several rORFs in the -RNA of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have the ability to suppress type-I interferon (IFN-I) production and facilitate vesicular stomatitis virus (VSV) infection. Additionally, we provide evidence that TuMV might utilize internal ribosome entry sites to translate these small rORFs. Our findings reveal that the -RNA of +ssRNA viruses can also have coding capacity and that small proteins encoded therein play critical roles in viral infection, unveiling a larger viral proteome than previously thought.

Keywords: Positive-sense single-stranded RNA viruses; negative-sense RNA; severe acute respiratory syndrome coronavirus 2; small proteins; turnip mosaic virus

DOI: https://doi.org/10.1016/j.molp.2023.09.020

Trends Biochem Sci. 2023 Sep 28. pii: S0968-0004(23)00228-1. [Epub ahead of print]

Two for one: regulatory RNAs that encode small proteins.

Jordan J Aoyama, Gisela Storz.

RNAs are commonly categorized as being either protein-coding mRNAs or noncoding RNAs. However, an increasing number of transcripts, in organisms ranging from bacteria to humans, are being found to have both coding and noncoding functions. In some cases, the sequences encoding the protein and the regulatory RNA functions are separated, while in other cases the sequences overlap. The protein and RNA can regulate similar or distinct pathways. Here we describe examples illustrating how these dual-function (also denoted bifunctional or dual-component) RNAs are identified and their mechanisms of action and cellular roles. We also discuss the synergy or competition between coding and RNA activity and how these regulators evolved, as well as how more dual-function RNAs might be discovered and exploited.

Keywords: bifunctional; dual component; dual function; miPEP; microprotein; small protein

DOI: https://doi.org/10.1016/j.tibs.2023.09.002