The RefSeq project at the National Center for Biotechnology Information (NCBI) maintains and curates a publicly available database of annotated genomic, transcript, and protein sequence records (http://www.ncbi.nlm.nih.gov/refseq/). The RefSeq project leverages the data submitted to the International Nucleotide Sequence Database Collaboration (INSDC) against a combination of computation, manual curation, and collaboration to produce a standard set of stable, non-redundant reference sequences. The RefSeq project augments these reference sequences with current knowledge including publications, functional features and informative nomenclature. The database currently represents sequences from more than 55,000 organisms (>4800 viruses, >40,000 prokaryotes and >10,000 eukaryotes; RefSeq release 71), ranging from a single record to complete genomes. This paper summarizes the current status of the viral, prokaryotic, and eukaryotic branches of the RefSeq project, reports on improvements to data access and details efforts to further expand the taxonomic representation of the collection. We also highlight diverse functional curation initiatives that support multiple uses of RefSeq data including taxonomic validation, genome annotation, comparative genomics, and clinical testing. We summarize our approach to utilizing available RNA-Seq and other data types in our manual curation process for vertebrate, plant, and other species, and describe a new direction for prokaryotic genomes and protein name management.

Reference sequence (RefSeq) database at NCBI: current status, taxonomic expansion, and functional annotation

Nonsense-mediated mRNA decay (NMD) is probably the best characterized eukaryotic RNA degradation pathway. Through intricate steps, a set of NMD factors recognize and degrade mRNAs with translation termination codons that are positioned in abnormal contexts. However, NMD is not only part of a general cellular quality control system that prevents the production of aberrant proteins. Mammalian cells also depend on NMD to dynamically adjust their transcriptomes and their proteomes to varying physiological conditions. In this Review, we discuss how NMD targets mRNAs, the types of mRNAs that are targeted, and the roles of NMD in cellular stress, differentiation and maturation processes.

/pdf/nonsense-mediated-mrna-decay-an-intricate-machinery-that-56a1ewt1zq.pdf

Nonsense-mediated mRNA decay: an intricate machinery that shapes transcriptomes.

Identification of the coding elements in the genome is a fundamental step to understanding the building blocks of living systems. Short peptides (< 100 aa) have emerged as important regulators of development and physiology, but their identification has been limited by their size. We have leveraged the periodicity of ribosome movement on the mRNA to define actively translated ORFs by ribosome footprinting. This approach identifies several hundred translated small ORFs in zebrafish and human. Computational prediction of small ORFs from codon conservation patterns corroborates and extends these findings and identifies conserved sequences in zebrafish and human, suggesting functional peptide products (micropeptides). These results identify micropeptide-encoding genes in vertebrates, providing an entry point to define their function in vivo.

https://www.embopress.org/doi/epdf/10.1002/embj.201488411

Identification of small ORFs in vertebrates using ribosome footprinting and evolutionary conservation

Although most mRNA molecules derived from protein-coding genes are destined to be translated into functional polypeptides, some are eliminated by cellular quality control pathways that collectively perform the task of mRNA surveillance. In the nonsense-mediated decay (NMD) pathway premature translation termination promotes the recruitment of a set of factors that destabilize a targeted mRNA. The same factors also seem to have key roles in repressing the translation of the mRNA, dissociating its terminating ribosome and messenger ribonucleoproteins (mRNPs), promoting the degradation of its truncated polypeptide product and possibly even feeding back to the site of transcription to interfere with splicing of the primary transcript.

NMD: a multifaceted response to premature translational termination.

The endoplasmic reticulum (ER) is a membranous intracellular organelle and the first compartment of the secretory pathway As such, the ER contributes to the production and folding of approximately one-third of cellular proteins, and is thus inextricably linked to the maintenance of cellular homeostasis and the fine balance between health and disease Specific ER stress signalling pathways, collectively known as the unfolded protein response (UPR), are required for maintaining ER homeostasis The UPR is triggered when ER protein folding capacity is overwhelmed by cellular demand and the UPR initially aims to restore ER homeostasis and normal cellular functions However, if this fails, then the UPR triggers cell death In this review, we provide a UPR signalling-centric view of ER functions, from the ER's discovery to the latest advancements in the understanding of ER and UPR biology Our review provides a synthesis of intracellular ER signalling revolving around proteostasis and the UPR, its impact on other organelles and cellular behaviour, its multifaceted and dynamic response to stress and its role in physiology, before finally exploring the potential exploitation of this knowledge to tackle unresolved biological questions and address unmet biomedical needs Thus, we provide an integrated and global view of existing literature on ER signalling pathways and their use for therapeutic purposes

/pdf/endoplasmic-reticulum-stress-signalling-from-basic-1ncpnvizt7.pdf

Endoplasmic Reticulum Stress signalling - from basic mechanisms to clinical applications

Upstream open reading frames (uORFs) are major gene expression regulatory elements. In many eukaryotic mRNAs, one or more uORFs precede the initiation codon of the main coding region. Indeed, several studies have revealed that almost half of human transcripts present uORFs. Very interesting examples have shown that these uORFs can impact gene expression of the downstream main ORF by triggering mRNA decay or by regulating translation. Also, evidence from recent genetic and bioinformatic studies implicates disturbed uORF-mediated translational control in the etiology of many human diseases, including malignancies, metabolic or neurologic disorders, and inherited syndromes. In this review, we will briefly present the mechanisms through which uORFs regulate gene expression and how they can impact on the organism's response to different cell stress conditions. Then, we will emphasize the importance of these structures by illustrating, with specific examples, how disturbed uORF-mediated translational control can be involved in the etiology of human diseases, giving special importance to genotype-phenotype correlations. Identifying and studying more cases of uORF-altering mutations will help us to understand and establish genotype-phenotype associations, leading to advancements in diagnosis, prognosis, and treatment of many human disorders.

/pdf/gene-expression-regulation-by-upstream-open-reading-frames-2k7f7kh3rv.pdf

Gene expression regulation by upstream open reading frames and human disease.

Nonsense-mediated mRNA decay (NMD) is a surveillance pathway that recognizes and rapidly degrades mRNAs containing premature termination codons (PTC). The strength of the NMD response appears to reflect multiple determinants on a target mRNA. We have previously reported that mRNAs containing PTCs in close proximity to the translation initiation codon (AUG-proximal PTCs) can substantially evade NMD. Here, we explore the mechanistic basis for this NMD resistance. We demonstrate that translation termination at an AUG-proximal PTC lacks the ribosome stalling that is evident in an NMD-sensitive PTC. This difference is associated with demonstrated interactions of the cytoplasmic poly(A)-binding protein 1, PABPC1, with the cap-binding complex subunit, eIF4G and the 40S recruitment factor eIF3 as well as the ribosome release factor, eRF3. These interactions, in combination, underlie critical 3'-5' linkage of translation initiation with efficient termination at the AUG-proximal PTC and contribute to an NMD-resistant PTC definition at an early phase of translation elongation.

https://repositorio.insa.pt/bitstream/10400.18/334/3/Interaction%20of%20PABPC1%20with%20the%20translation%20initiation.pdf

Interaction of PABPC1 with the translation initiation complex is critical to the NMD resistance of AUG-proximal nonsense mutations

Messenger RNA (mRNA) stability is a critical determinant that affects gene expression. Many pathways have evolved to modulate mRNA stability in response to developmental, physiological and/or environmental stimuli. Eukaryotic mRNAs have a considerable range of half-lives, from as short as a few minutes to as long as several days. Human globin mRNAs constitute an example of highly stable mRNAs. However, a wide variety of naturally occurring mutations that result in the clinical syndrome of thalassemia can trigger accelerated mRNA decay thus controlling mRNA quality prior to translation. Distinct surveillance mechanisms have been described as being targeted for specific defective globin mRNAs. Here, we review mRNA stability mechanisms implicated in the control of β-globin gene expression and the surveillance pathways that prevent translation of aberrant β-globin mRNAs. In addition, we emphasize the importance of these pathways in modulating the severity of the β-thalassemia phenotype.

https://repositorio.insa.pt/bitstream/10400.18/333/1/I%20Peixeiro%20et%20al.%202011.pdf

Control of human beta-globin mRNA stability and its impact on beta-thalassemia phenotype.

MPTP-induced dopaminergic neurotoxicity involves major biochemical processes such as oxidative stress and impaired energy metabolism, leading to a significant reduction in the number of nigrostriatal dopaminergic neurons. Glutathione S-transferase pi (GSTpi) is a phase II detoxifying enzyme that provides protection of cells from injury by toxic chemicals and products of oxidative stress. In humans, polymorphisms of GSTP1 affect substrate selectivity and stability increasing the susceptibility to parkinsonism-inducing effects of environmental toxins. Given the ability of MPTP to increase the levels of reactive oxygen species and the link between altered redox potential and the expression and activity of GSTpi, we investigated the effect of MPTP on GSTpi cellular concentration in an in vivo model of Parkinson’s disease. The present study demonstrates that GSTpi is actively expressed in both substantia nigra pars compacta and striatum of C57BL/6 mice brain, mostly in oligodendrocytes and astrocytes. After systemic administration of MPTP, GSTpi expression is significantly increased in glial cells in the vicinity of dopaminergic neurons cell bodies and fibers. The results suggest that GSTpi expression may be part of the mechanism underlying the ability of glial cells to elicit protection against the mechanisms involved in MPTP-induced neuronal death.

GSTpi Expression in MPTP-Induced Dopaminergic Neurodegeneration of C57BL/6 Mouse Midbrain and Striatum

https://repositorio.insa.pt/bitstream/10400.18/1042/3/Abstract%20FEBS%20-%20Algarve%202012.pdf

Nonsense-mediated decay resistance of AUG-proximal nonsense-mutated transcripts relies on the interaction of PABPC1 with the translation initiation complex

Isabel Peixeiro

Papers

Gene expression regulation by upstream open reading frames and human disease.

Interaction of PABPC1 with the translation initiation complex is critical to the NMD resistance of AUG-proximal nonsense mutations

Control of human beta-globin mRNA stability and its impact on beta-thalassemia phenotype.

GSTpi Expression in MPTP-Induced Dopaminergic Neurodegeneration of C57BL/6 Mouse Midbrain and Striatum

Nonsense-mediated decay resistance of AUG-proximal nonsense-mutated transcripts relies on the interaction of PABPC1 with the translation initiation complex