Genes within Genes in Bacterial Genomes

Abstract

Genetic coding in bacteria largely operates via the "one gene-one protein" paradigm. However, the peculiarities of the mRNA structure, the versatility of the genetic code, and the dynamic nature of translation sometimes allow organisms to deviate from the standard rules of protein encoding. Bacteria can use several unorthodox modes of translation to express more than one protein from a single mRNA cistron. One such alternative path is the use of additional translation initiation sites within the gene. Proteins whose translation is initiated at different start sites within the same reading frame will differ in their N termini but will have identical C-terminal segments. On the other hand, alternative initiation of translation in a register different from the frame dictated by the primary start codon will yield a protein whose sequence is entirely different from the one encoded in the main frame. The use of internal mRNA codons as translation start sites is controlled by the nucleotide sequence and the mRNA folding. The proteins of the alternative proteome generated via the "genes-within-genes" strategy may carry important functions. In this review, we summarize the currently known examples of bacterial genes encoding more than one protein due to the utilization of additional translation start sites and discuss the known or proposed functions of the alternative polypeptides in relation to the main protein product of the gene. We also discuss recent proteome- and genome-wide approaches that will allow the discovery of novel translation initiation sites in a systematic fashion.

Diverse CRISPR-Cas Complexes Require Independent Translation of Small and Large Subunits from a Single Gene

“…Furthermore, mutations within the RBS and/or start codon, or a scrambled control, abolished expression (Figure 2B). Generation of alternative proteins from internal translation represents a cryptic part of the bacterial proteome that is now just being revealed (Meydan et al, 2018(Meydan et al, , 2019. The expression of Cas11d from within cas10d transcripts is the first known report of alternative internal translation in CRISPR-Cas systems.…”

Complementary Ribo-seq approaches map the translatome and provide a small protein census in the foodborne pathogenCampylobacter jejuni

“…While internal TIS are well known to generate shorter, functional proteins (Meydan et al, 2018), 3′ end truncations are less well-characterized. C-terminal truncations might arise via, e.g., point mutations, alternative decoding, or ribosomal frameshifting.…”

“…This includes hundreds of potential small non-coding regulatory RNAs (sRNAs) in intergenic regions, which act as key regulators of bacterial stress responses, fitness, antibiotic resistance, and virulence (Felden and Augagneur, 2021;Hör et al, 2020;Svensson and Sharma, 2016;Westermann, 2018). More recently, the concept of genes within genes has emerged in different forms, e.g., as sRNAs derived from mRNA untranslated regions (Ponath et al, 2022), dual-function (coding and regulatory) sRNAs (Raina et al, 2018), or alternative open reading frames (ORFs) generated by internal start codons (Adams and Storz, 2020;Meydan et al, 2018;Orr et al, 2020). Therefore, bacterial genomes likely still hide genes with key roles in physiology and/or virulence.…”

“…More recently, the concept of genes within genes has emerged in different forms, e . g ., as sRNAs derived from mRNA untranslated regions (Ponath et al, 2022), dual-function (coding and regulatory) sRNAs (Raina et al, 2018), or alternative open reading frames (ORFs) generated by internal start codons (Adams and Storz, 2020; Meydan et al, 2018; Orr et al, 2020). Therefore, bacterial genomes likely still hide genes with key roles in physiology and/or virulence.…”

Pervasive translation in Mycobacterium tuberculosis

“…Ribo-RET is a modified form of Ribo-seq in which bacterial cells are treated with the antibiotic retapamulin before lysis; retapamulin traps bacterial ribosomes at sites of translation initiation, whereas elongating ribosomes are free to complete translation ( Meydan et al, 2019 ). Thus, Ribo-RET facilitates the identification of overlapping ORFs by limiting the signal to the start codons ( Meydan et al, 2018 ; Meydan et al, 2019 ). Ribo-RET was recently applied to E. coli , revealing start codons for many previously undescribed ORFs ( Meydan et al, 2019 ; Stringer et al, 2021 ; Weaver et al, 2019 ), including sORFs, and ORFs positioned in frame with annotated ORFs, such that the translated protein is an isoform of the previously described protein.…”