Purriato is a conserved small open reading frame gene that interacts with the CASA pathway to regulate muscle homeostasis and epithelial tissue growth in Drosophila.

Recent advances in proteogenomic techniques and bioinformatic pipelines have permitted the detection of thousands of translated small Open Reading Frames (smORFs), which contain less than 100 codons, in eukaryotic genomes. Hundreds of these actively translated smORFs display conserved sequence, structure and evolutionary signatures indicating that the translated peptides could fulfil important biological roles. Despite their abundance, only tens of smORF genes have been fully characterised; these act mainly as regulators of canonical proteins involved in essential cellular processes. Importantly, some of these smORFs display conserved functions with their mutations being associated with pathogenesis. Thus, investigating smORF roles in Drosophila will not only expand our understanding of their functions but it may have an impact in human health. Here we describe the function of a novel and essential Drosophila smORF gene named purriato (prto). prto belongs to an ancient gene family whose members have expanded throughout the Protostomia clade. prto encodes a transmembrane peptide which is localized in endo-lysosomes and perinuclear and plasma membranes. prto is dynamically expressed in mesodermal tissues and imaginal discs. Targeted prto knockdown (KD) in these organs results in changes in nuclear morphology and endo-lysosomal distributions correlating with the loss of sarcomeric homeostasis in muscles and reduction of mitosis in wing discs. Consequently, prto KD mutants display severe reduction of motility, and shorter wings. Finally, our genetic interaction experiments show that prto function is closely associated to the CASA pathway, a conserved mechanism involved in turnover of mis-folded proteins and linked to muscle dystrophies and neurodegenerative diseases. Thus, this study shows the relevance of smORFs in regulating important cellular functions and supports the systematic characterisation of this class of genes to understand their functions and evolution.

Translation and natural selection of micropeptides from long non-canonical RNAs.

Long noncoding RNAs (lncRNAs) are transcripts longer than 200 nucleotides but lacking canonical coding sequences. Apparently unable to produce peptides, lncRNA function seems to rely only on RNA expression, sequence and structure. Here, we exhaustively detect in-vivo translation of small open reading frames (small ORFs) within lncRNAs using Ribosomal profiling during Drosophila melanogaster embryogenesis. We show that around 30% of lncRNAs contain small ORFs engaged by ribosomes, leading to regulated translation of 100 to 300 micropeptides. We identify lncRNA features that favour translation, such as cistronicity, Kozak sequences, and conservation. For the latter, we develop a bioinformatics pipeline to detect small ORF homologues, and reveal evidence of natural selection favouring the conservation of micropeptide sequence and function across evolution. Our results expand the repertoire of lncRNA biochemical functions, and suggest that lncRNAs give rise to novel coding genes throughout evolution. Since most lncRNAs contain small ORFs with as yet unknown translation potential, we propose to rename them "long non-canonical RNAs".

Pegasus, a small extracellular peptide enhancing short-range diffusion of Wingless.

Small Open Reading Frames (smORFs) coding for peptides of less than 100 amino-acids are an enigmatic and pervasive gene class, found in the tens of thousands in metazoan genomes. Here we reveal a short 80 amino-acid peptide (Pegasus) which enhances Wingless/Wnt1 protein short-range diffusion and signalling. During Drosophila wing development, Wingless has sequential functions, including late induction of proneural gene expression and wing margin development. Pegasus mutants produce wing margin defects and proneural expression loss similar to those of Wingless. Pegasus is secreted, and co-localizes and co-immunoprecipitates with Wingless, suggesting their physical interaction. Finally, measurements of fixed and in-vivo Wingless gradients support that Pegasus increases Wingless diffusion in order to enhance its signalling. Our results unveil a new element in Wingless signalling and clarify the patterning role of Wingless diffusion, while corroborating the link between small open reading frame peptides, and regulation of known proteins with membrane-related functions.

New Peptides Under the s(ORF)ace of the Genome.

Hundreds, perhaps thousands of previously unidentified functional small peptides could exist in most genomes, but these sequences have been generally overlooked. The discovery of genes encoding small peptides with important functions in different organisms has ignited the interest in these sequences and led to an increasing amount of effort towards their identification. Here, we review the advances, both computational and biochemical, that are leading the way in the discovery of putatively functional small open reading frame genes (smORFs), as well as the functional studies that have been carried out as a consequence of these searches. The evidence suggests that smORFs form a substantial part of our genomes, and that their encoded peptides could have a variety of important cellular functions.

Hemotin, a Regulator of Phagocytosis Encoded by a Small ORF and Conserved across Metazoans.

Translation of hundreds of small ORFs (smORFs) of less than 100 amino acids has recently been revealed in vertebrates and Drosophila. Some of these peptides have essential and conserved cellular functions. In Drosophila, we have predicted a particular smORF class encoding ~80 aa hydrophobic peptides, which may function in membranes and cell organelles. Here, we characterise hemotin, a gene encoding an 88aa transmembrane smORF peptide localised to early endosomes in Drosophila macrophages. hemotin regulates endosomal maturation during phagocytosis by repressing the cooperation of 14-3-3ζ with specific phosphatidylinositol (PI) enzymes. hemotin mutants accumulate undigested phagocytic material inside enlarged endo-lysosomes and as a result, hemotin mutants have reduced ability to fight bacteria, and hence, have severely reduced life span and resistance to infections. We identify Stannin, a peptide involved in organometallic toxicity, as the Hemotin functional homologue in vertebrates, showing that this novel regulator of phagocytic processing is widely conserved, emphasizing the significance of smORF peptides in cell biology and disease.

Enfermedad de Hirschsprung, mutación del protooncogén RET y tiroidectomía profiláctica / Hirschsprung's disease, RET proto-oncogene mutation and prophylactic thyroidectomy

No disponible

Tarsal-less peptides control Notch signalling through the Shavenbaby transcription factor.

The formation of signalling boundaries is one of the strategies employed by the Notch (N) pathway to give rise to two distinct signalling populations of cells. Unravelling the mechanisms involved in the regulation of these signalling boundaries is essential to understanding the role of N during development and diseases. The function of N in the segmentation of the Drosophila leg provides a good system to pursue these mechanisms at the molecular level. Transcriptional and post-transcriptional regulation of the N ligands, Serrate (Ser) and Delta (Dl) generates a signalling boundary that allows the directional activation of N in the distalmost part of the segment, the presumptive joint. A negative feedback loop between odd-skipped-related genes and the N pathway maintains this signalling boundary throughout development in the true joints. However, the mechanisms controlling N signalling boundaries in the tarsal joints are unknown. Here we show that the non-canonical tarsal-less (tal) gene (also known as pri), which encodes for four small related peptides, is expressed in the N-activated region and required for joint development in the tarsi during pupal development. This function of tal is both temporally and functionally separate from the tal-mediated tarsal intercalation during mid-third instar that we reported previously. In the pupal function described here, N signalling activates tal expression and reciprocally Tal peptides feedback on N by repressing the transcription of Dl in the tarsal joints. This Tal-induced repression of Dl is mediated by the post-transcriptional activation of the Shavenbaby transcription factor, in a similar manner as it has been recently described in the embryo. Thus, a negative feedback loop involving Tal regulates the formation and maintenance of a Dl+/Dl- boundary in the tarsal segments highlighting an ancient mechanism for the regulation of N signalling based on the action of small cell signalling peptides.

Comparative analysis of leg and antenna development in wild-type and homeotic Drosophila melanogaster.

The insect leg and antenna are thought to be homologous structures, evolved from a common ancestral appendage. The homeotic transformations of antenna to leg in Drosophila produced by mutation of the Hox gene Antennapedia are position-specific, such that every particular antenna structure is transformed into a specific leg counterpart. This has been taken to suggest that the developmental programmes of these two appendages are still similar. In particular, the mechanisms for the specification of a cell's position within the appendage would be identical, only their interpretation would be different and subject to homeotic gene control. Here we explore the degree of conservation between the developmental programmes of leg and antenna in Drosophila and other dipterans, in wild-type and homeotic conditions. Most of the appendage pattern-forming genes are active in both appendages, and their expression domains are partially conserved. However, the regulatory relationships and interactions between these genes are different, and in fact cells change their expression while undergoing homeotic transformation. Thus, the positional information, and the mechanisms which generate it, are not strictly conserved between leg and antenna; and homeotic genes alter the establishment of positional clues, not only their interpretation. The partial conservation of pattern-forming genes in both appendages ensures a predictable re-specification of positional clues, producing the observed positional specificity of homeotic transformations.