Abstract
Genes encoding small open-reading frames (smORFs) have been characterized as essential players of developmental processes. The smORF tarsaless/mille-pattes/polished-rice has been thoroughly investigated in holometabolous insects, such as the fruit fly Drosophila melanogaster and the red flour beetle Tribolium castaneum, while its function in hemimetabolous insects was only recently investigated. Thus, we analyzed the function of the tal/pri/mlpt ortholog in a hemimetabolous insect, the kissing bug Rhodnius prolixus (Rp). Sequence analysis shows that Rp-mlpt polycistronic mRNA encodes two small peptides (11 to 14 amino acids) containing a LDPTG motif. Interestingly, a new hemipteran-specific conserved peptide of approximately 80 amino acids was also identified by in silico analysis. In silico docking analysis supports the binding of the small Mlpt peptides to the transcription factor Shavenbaby. Rp-mlpt in situ hybridization and knockdown via RNA interference showed a conserved role of Rp-mlpt during embryogenesis, with a major role in the regulation of thoracic versus abdominal segmentation, leg development and head formation. Altogether, our study shows that mlpt segmentation role is conserved in the common ancestor of Paraneoptera and suggests that polycistronic genes might generate order specific smORFs.
Highlights
A large number of essential genes required for biological processes have been discovered by genetic screenings in model organisms such as the fruit fly Drosophila melanogaster (e.g., Nüsslein-Volhard and Wieschaus, 1980)
Black lines comprise the whole polycistronic mRNA containing the small open reading frames (smORFs) encoding peptides displayed in red, green, and pink boxes. (B) Alignment of the previously described small peptide containing an LDPTG domain (Savard et al, 2006; Galindo et al, 2007; Kondo et al, 2007), conserved in several arthropod species and whose number of paralogs in a single polycistronic gene varies among arthropod species. (C) Alignment of a hemipteran specific peptide of about 80 amino acids. (D) Alignment of triatomine conserved smORF encoded peptides. #indicates that translation is not interrupted in this sequence
Previous BLAST searches for genes encoding mlpt peptides against different arthropod genomes and transcriptomes provided evidence that this gene containing smORFs is restricted to insects and crustaceans (Galindo et al, 2007)
Summary
A large number of essential genes required for biological processes have been discovered by genetic screenings in model organisms such as the fruit fly Drosophila melanogaster (e.g., Nüsslein-Volhard and Wieschaus, 1980). While most loci important for developmental processes were identified in these original screenings, recent genetic and expression analyses in D. melanogaster and in the tal/mlpt/pri in the Hemiptera Rhodnius beetle Tribolium castaneum showed that genes previously classified as putative non-coding RNAs encode functional small open reading frames (smORFs) or sORFs (Pueyo et al, 2016a; Chekulaeva and Rajewsky, 2018). Experimental analysis of conserved smORFs such as sarcolamban (scl), a conserved peptide involved in Ca2+ uptake at the sarco-endoplasmic reticulum (Magny et al, 2013), and hemotin (Pueyo et al, 2016b), a conserved phagocytosis regulator, provided further evidence that genes containing smORFs might constitute a reservoir of important players in metazoan genomes. These “dwarf ” smORFs are generally shorter (around 20 amino-acid long), less conserved and mostly found in 5′-UTRs and noncoding RNAs
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