APGW/AKH Precursor from Rotifer Brachionus plicatilis and the DNA Loss Model Explain Evolutionary Trends of the Neuropeptide LWamide, APGWamide, RPCH, AKH, ACP, CRZ, and GnRH Families.

In the year 2002, DNA loss model (DNA-LM) postulated that neuropeptide genes to emerged through codons loss via the repair of damaged DNA from ancestral gene namely Neuropeptide Precursor Predictive (NPP), which organization correspond two or more neuropeptides precursors evolutive related. The DNA-LM was elaborated according to amino acids homology among LWamide, APGWamide, red pigment-concentrating hormone (RPCH), adipokinetic hormones (AKHs) and in silico APGW/RPCH NPPAPGW/AKH NPP were proposed. With the above principle, it was proposed the evolution of corazonin (CRZ), gonadotropin-releasing hormone (GnRH), AKH, and AKH/CRZ (ACP), but any NPP never was considered. However, the evolutive relation via DNA-LM among these neuropeptides precursors not has been established yet. Therefore, the transcriptomes from crabs Callinectes toxotes and Callinectes arcuatus were used to characterized ACP and partial CRZ precursors, respectively. BLAST alignment with APGW/RPCH NPP and APGW/AKH NPP allow identified similar NPP in the rotifer Brachionus plicatilis and other invertebrates. Moreover, three bioinformatics algorithms and manual verification were used to purify 13,778 sequences, generating a database with 719 neuropeptide precursors. Phylogenetic trees with the DNA-LM parameters showed that some ACP, CRZ, AKH2 and two NPP share nodes with GnRH from vertebrates and some of this neuropeptide had nodes in invertebrates. Whereas the phylogenetic tree with standard parameters do not showed previous node pattern. Robinson-Foulds metric corroborates the differences among phylogenetic trees. Homology relationship showed four putative orthogroups; AKH4, CRZ, and protostomes GnRH had individual group. This is the first demonstration of NPP in species and would explain the evolution neuropeptide families by the DNA-LM.

Chronic cough in post-COVID syndrome: Laryngeal electromyography findings in vagus nerve neuropathy.

BACKGROUND: Despite being a new entity, there is a large amount of information on the characteristics of SARS-CoV-2 infection and the symptoms of the acute phase; however, there are still many unknowns about the clinical features and pathophysiology of post-COVID syndrome. Refractory chronic cough is one of the most prevalent symptoms and carries both a medical problem and a social stigma. Many recent studies have highlighted the role of SARS-CoV-2 neurotropism, but no studies have demonstrated vagus nerve neuropathy as a cause of persistent chronic cough or other COVID-19 long-term effects. OBJECTIVE: The main objective was to assess the involvement of the vagus nerve neuropathy as a cause of chronic cough and other post-COVID syndrome symptoms. MATERIAL AND METHODS: This was a single-center observational study with prospective clinical data collected from 38 patients with chronic cough and post-COVID-19 syndrome. Clinical characteristics and laryngeal electromyographic findings were analyzed. RESULTS: Clinical data from 38 patients with chronic cough after 12 weeks of the acute phase of COVID-19 infection were analyzed. Of these patients, 81.6% suffered from other post-COVID conditions and, 73.6% reported fluctuating evolution of symptoms. Laryngeal electromyography (LEMG) of the thyroarytenoid (TA) muscles and cricothyroid (CT) muscles was pathological in 76.3% of the patients. Of the patients with abnormal LEMG, chronic denervation was the most frequent finding (82.8%), 10.3% presented acute denervation signs, and 6.9% presented myopathic pattern in LEMG. CONCLUSIONS: LEMG studies suggest the existence of postviral vagus nerve neuropathy after SARS-CoV-2 infection that could explain chronic cough in post-COVID syndrome.

Hallazgos electroencefalográficos compatibles con encefalopatía en pacientes ingresados por covid-19 / Electroencephalographic findings compatible with encephalopathy in patients admitted for COVID-19

RESUMEN INTRODUCCIÓN: La covid-19 afecta principalmente al aparato respiratorio, sin embargo, también se ha descrito afectación tanto directa como indirecta en el sistema nervioso central y periférico, lo cual ocasiona una gran variedad de manifestaciones neurológicas, siendo la encefalopatía una de las más frecuentemente observadas. OBJETIVO: Se busca mostrar la utilidad del video-electroencefalograma (vEEG) en el diagnóstico de encefalopatía en pacientes ingresados por covid-19, así como su valor para determinar el pronóstico de estos pacientes. MÉTODOS: Estudio observacional retrospectivo con 76 vEEG de 41 pacientes con covid-19 confirmada. Los estudios se han realizado entre los meses de marzo del 2020 y junio del 2021. Se estudió la gravedad de la enfermedad, así como sus características clínicas y neurológicas, el tratamiento farmacológico y los hallazgos electroencefalográficos según el grado de disfunción de la encefalopatía que desarrollaron estos pacientes. RESULTADOS: De los 41 pacientes, 12 (29 %) presentaron signos electroencefalográficos de disfunción cerebral leve, 15 (37 %) disfunción cerebral moderada y 14 (34 %) disfunción cerebral severa, los cuales se asociaron con una mayor mortalidad. CONCLUSIONES: En los 76 vEEG realizados a los 41 pacientes ingresados con encefalopatías asociadas con infección por covid-19, no se observó un patrón distinto a los descritos en encefalopatías de otras etiologías. El vEEG fue útil para confirmar la sospecha clínica de una disfunción cerebral en pacientes con encefalopatías asociadas con infección por covid-19 y para asignarle un grado de severidad, confirmando su beneficio como biomarcador diagnóstico y pronóstico.

ABSTRACT INTRODUCTION: COVID-19 mainly affects the respiratory system; however, both direct and indirect involvement of the central and peripheral nervous system has also been described, causing a wide variety of neurological manifestations, with encephalopathy being one of the most frequently observed neurological manifestations. OBJECTIVE: With this article we intend to show the usefulness of vEEG in the diagnosis of encephalopathy in patients referred for COVID-19 who develop this neurological complication, as well as its value in determining the prognosis of these patients. METHODS: Retrospective observational study with 76 video-electroencephalograms of 41 patients with confirmed COVID-19 infection. The studies were performed during the months of March 2020 through June 2021. Disease severity, clinical and neurological features, pharmacological treatment and electroencephalographic indings were studied according to the degree of encephalopathy dysfunction these patients developed. RESULTS: Of the 41 patients, 12 (29 %) presented electroencephalographic signs of mild cerebral dysfunction, 15 (37 %) moderate cerebral dysfunction, and 14 (34 %) severe cerebral dysfunction, which were associated with higher mortality. CONCLUSIONS: In the 76 vEEG performed in the 41 patients admitted with encephalopathies associated with COVID-19 infection, no pattern different from that described in encephalopathies of other etiologies was observed. The vEEG was useful to confirm the clinical suspicion of brain dysfunction in patients with encephalopathies associated with COVID-19 infection and to assign a degree of severity, confirming its benefit as a diagnostic and prognostic biomarker.

APLICACIÓN DEL MODELO DE PÉRDIDA DE ADN PARA EL DISEÑO DE CEBADORES EN Potamotrygon magdalenae (POTAMOTRYGONIDAE) / Application of the model of DNA loss for desing of primers in Potamotrygon magdalenae (Potamotrygonidae)

RESUMEN El diseño de cebadores es fundamental para amplificar regiones de genes debido a que la especificidad que mantienen cebador-secuencia de interés puede causar el éxito o fracaso en la reacción de PCR. En relación a Potamotrygon magdalenae (especie de interés de acuerdo al PAN Tiburones-Colombia), existe poca información disponible de aspectos relacionados con la genética poblacional de esta Raya. El objetivo del presente trabajo consistió en diseñar cebadores bajo los criterios del Modelo de Pérdida de ADN (DNA-LM), que permitan evaluar el estado genético de las poblaciones de P. magdalenae. Alineamos secuencias de la superfamilia Dayastoidea, disponibles en el NCBI, de los genes mitocondriales Citocromo C Oxidasa 1 (MT-CO1) y Citocromo b (MT-CYB). Seguimos los parámetros Gap open penalty (5), Gap extension penalti (0,2) y Terminalgap penalties (0,1) y seleccionamos dos pares de cebadores de acuerdo con el DNA-LM. Estimamos el producto amplificado del gen MT-CO1 en 916 pb y del gen MT-CYB en 774 pb, en muestras de P. magdalenae procedentes de diferentes ciénagas del Magdalena medio. Discutimos los resultados desde la perspectiva de validar la especificidad de los cebadores diseñados, teniendo en cuenta la correspondencia e identidad de las secuencias de los genes considerados. Los cebadores aquí reportados pueden contribuir a ampliar el conocimiento de la genética poblacional, biogeografía y filogenética de la raya de agua dulce P. magdalenae.

ABSTRACT The primer design is critical for amplifying gene regions because the specificity that primer-sequence maintain can cause success or failure in the PCR reaction. Concerning Potamotrygon magdalenae (a species of interest according to the PAN Tiburones-Colombia), there is little information available about aspects related to population genetics of this river stingray. This study aimed to design primers under DNA Loss Model (DNA-LM) criteria, which will allow assessing the genetics status of populations of P. magdalenae. We aligned sequences from the superfamily Dayastoidea, available in the NCBI, of the mitochondrial genes Cytochrome C Oxidase 1 (MT-CO1) and Cytochromeb (MT-CYB). We followed the parameters Gap open penalty (5), Gap extension penalty (0.2) and Terminal gap penalties (0.1) and selected two pairs of primers according to the DNA-LM. We calculated the amplified product of the MT-CO1 gene in 916 pb and the MT-CYB gene in 774 pb in samples from different swamps of the middle Magdalena. We discussed the results from the perspective of validating the specificity of the primers designed, considering the correspondence and identity of gene sequences considered. The primers reported here will contribute to broadening the knowledge of population genetics, biogeography and phylogenetic of the river stingray P. magdalenae.

Comparison between cultured and wild Pacific white shrimp (Penaeus vannamei) vitellogenesis: next-generation sequencing and relative expression of genes directly and indirectly related to reproduction.

Shrimp fisheries are among the most important fisheries worldwide, and shrimp culture has increased considerably in recent years. Most current studies on reproduction-related genes have been conducted on cultured shrimp. However, gene expression is intimately linked to physiological and environmental conditions, and therefore an organism's growth environment has a great influence on reproduction. Thus, gene expression profiling, should be applied in fisheries studies. Here, we identified the expression patterns of 76 reproduction-related genes in P. vannamei via the analysis of pooled transcriptomes from a time-series experiment encompassing a full circadian cycle. The expression patterns of genes associated both directly (Vtg, ODP, and ProR) and indirectly (FAMet, CruA1, and CruC1) with reproduction were evaluated, as these genes could be used as molecular markers of previtellogenic and vitellogenic maturation stages. The evaluated genes were prominently upregulated during vitellogenic stages, with specific expression patterns depending on the organism's environment, diet, and season. Vtg, ProR, ODP, and FaMet could serve as molecular markers for both wild and cultured organisms.

Reproduction and community structure of fish from winter catch sites from industrial shrimp bycatch from the northeast and southeast Mexican Pacific.

The shrimp fishery is one of the most important fisheries in the world, although the low selectivity from trawling nets has led to the capture of a large number of non-target species. Shrimp-bycatch species include a large number of fish and invertebrate species, of which fish species are the most abundant. The present study aims to determine the community structure as well as the average sizes at first maturity of the fish species from shrimp-bycatch caught from industrial fisheries in the Mexican Pacific from Sinaloa to Guerrero, from January to March 2015. The shrimp-bycatch fish diversity value was found to be 2.22. A total of 37 species of finfish were found, of which five were considered rare. The fish species with the highest Importance Value Index (IVI) levels were Pseudupeneus grandisquamis, Paralichthys woolmani, Lutjanus peru and Diapterus peruvianus. The average size at first maturity was calculated for all species. Of the analysed organisms, 90% were in the juvenile stage, including species with riverine and artisanal fisheries. The present study demonstrates the risk within marine populations to different non-target species due to the poor selectivity of shrimp trawls.

Mycobacterium tuberculosis as a cause of mandibular osteomyelitis in a young woman: a case report.

BACKGROUND: Tuberculosis is considered an emerging disease worldwide; in the last 10 years, its incidence has increased to more than 9.6 million cases of active tuberculosis. In 2014, it resulted in 1.5 million patient deaths. However, oral presentation with bone involvement occurs in less than 3% of all reported cases and rarely arouses clinical suspicion on initial presentation. CASE PRESENTATION: A 15-year-old Mexican girl who had a previous diagnosis of neurofibromatosis presented to our hospital with pain and swelling in the region of the left mandibular body since November 2011. A clinical examination revealed pain in the mandibular region, a mass of soft consistency that seemed to involve bone, and a fistula with discharge of intraoral purulent material. Additionally, tachycardia and hyperthermia were observed. The left submental and submandibular regions had a 12-cm-diameter swelling, which was well-delineated and nonerythematous. The final diagnosis was established by real-time polymerase chain reaction. CONCLUSIONS: The final diagnosis of rare cases of tuberculous osteomyelitis in the jaw can be established by deoxyribonucleic acid (DNA) identification of Mycobacterium tuberculosis in the lesion. Simple and fast complementary diagnosis by real-time polymerase chain reaction is a fundamental approach to establishing early and effective pharmacological and surgical treatment.

Analysis of chemokines and receptors expression profile in the myelin mutant taiep rat.

Taiep rat has a failure in myelination and remyelination processes leading to a state of hypomyelination throughout its life. Chemokines, which are known to play a role in inflammation, are also involved in the remyelination process. We aimed to demonstrate that remyelination-stimulating factors are altered in the brainstem of 1- and 6-month-old taiep rats. We used a Rat RT(2) Profiler PCR Array to assess mRNA expression of 84 genes coding for cytokines, chemokines, and their receptors. We also evaluated protein levels of CCL2, CCR1, CCR2, CCL5, CCR5, CCR8, CXCL1, CXCR2, CXCR4, FGF2, and VEGFA by ELISA. Sprague-Dawley rats were used as a control. PCR Array procedure showed that proinflammatory cytokines were not upregulated in the taiep rat. In contrast, some mRNA levels of beta and alpha chemokines were upregulated in 1-month-old rats, but CXCR4 was downregulated at their 6 months of age. ELISA results showed that CXCL1, CCL2, CCR2, CCR5, CCR8, and CXCR4 protein levels were decreased in brainstem at the age of 6 months. These results suggest the presence of a chronic neuroinflammation process with deficiency of remyelination-stimulating factors (CXCL1, CXCR2, and CXCR4), which might account for the demyelination in the taiep rat.

A novel strategy to isolate cell-envelope mutants resistant to phage infection: bacteriophage mEp213 requires lipopolysaccharides in addition to FhuA to enter Escherichia coli K-12.

We have developed a direct and efficient strategy, based on a three-step method, to select bacterial cell-envelope mutants resistant to bacteriophage infection. Escherichia coli K-12 strain W3110 underwent classical transposon mutagenesis followed by replica plating and selection for mutants resistant to infection by coliphage mEp213. To verify that phage resistance was due to mutations in the cell envelope, we transformed host cells with the viral genome using electroporation and selected those in which virions were subsequently detected in the supernatant. Among the nine mutants resistant to coliphage infection that we selected, six were in the fhuA gene, two were mutated in the waaC gene, and one was mutated in the gmhD gene. The latter two gene products are involved in the synthesis of lipopolysaccharide (LPS). The efficiency of plating and adsorption of phage mEp213 was affected in these mutants. We verified that LPS is required for the efficient infection of phage λ as well. We propose that this mutation-and-selection strategy can be used to find host factors involved in the initial steps of phage infection for any cognate pair of phage and bacteria.

Activity inference for Ambient Intelligence through handling artifacts in a healthcare environment.

Human activity inference is not a simple process due to distinct ways of performing it. Our proposal presents the SCAN framework for activity inference. SCAN is divided into three modules: (1) artifact recognition, (2) activity inference, and (3) activity representation, integrating three important elements of Ambient Intelligence (AmI) (artifact-behavior modeling, event interpretation and context extraction). The framework extends the roaming beat (RB) concept by obtaining the representation using three kinds of technologies for activity inference. The RB is based on both analysis and recognition from artifact behavior for activity inference. A practical case is shown in a nursing home where a system affording 91.35% effectiveness was implemented in situ. Three examples are shown using RB representation for activity representation. Framework description, RB description and CALog system overcome distinct problems such as the feasibility to implement AmI systems, and to show the feasibility for accomplishing the challenges related to activity recognition based on artifact recognition. We discuss how the use of RBs might positively impact the problems faced by designers and developers for recovering information in an easier manner and thus they can develop tools focused on the user.

Influence of codon usage bias on FGLamide-allatostatin mRNA secondary structure.

The FGLamide allatostatins (ASTs) are invertebrate neuropeptides which inhibit juvenile hormone biosynthesis in Dictyoptera and related orders. They also show myomodulatory activity. FGLamide AST nucleotide frequencies and codon bias were investigated with respect to possible effects on mRNA secondary structure. 367 putative FGLamide ASTs and their potential endoproteolytic cleavage sites were identified from 40 species of crustaceans, chelicerates and insects. Among these, 55% comprised only 11 amino acids. An FGLamide AST consensus was identified to be (X)(1→16)Y(S/A/N/G)FGLGKR, with a strong bias for the codons UUU encoding for Phe and AAA for Lys, which can form strong Watson-Crick pairing in all peptides analyzed. The physical distance between these codons favor a loop structure from Ser/Ala-Phe to Lys-Arg. Other loop and hairpin loops were also inferred from the codon frequencies in the N-terminal motif, and the first amino acids from the C-terminal motif, or the dibasic potential endoproteolytic cleavage site. Our results indicate that nucleotide frequencies and codon usage bias in FGLamide ASTs tend to favor mRNA folds in the codon sequence in the C-terminal active peptide core and at the dibasic potential endoproteolytic cleavage site.

Purification of bacterial genomic DNA in less than 20 min using chelex-100 microwave: examples from strains of lactic acid bacteria isolated from soil samples.

We established a Chelex 100-Microwave method for the purification of bacterial genomic DNA (gDNA) in less than 20 min with high yield and good quality, useful for multiple purposes. It combines Chelex 100, proteinase K, RNase A and heating in a microwave oven. The resulting gDNA was used directly to identify bacterial species of the Order Lactobacillales by means of PCR amplification of their 16S rDNA gene, isolated from sediments on the Yucatan Peninsula, Mexico. This method produced gDNA free of phenolic and protein residual contaminants from 100 of these isolated bacteria. 16S rDNA amplification and sequencing showed Pediococcus acidilactici to prevail in inland lagoons, and Pediococcus pentosaceus, Lactobacillus plantarum, Lactobacillus sp., and Lactobacillus fermentum to be most abundant in the soils of livestock farms. The combination of Chelex 100, enzymes and microwave heating used in the Chelex 100-Microwave method produced large amounts of highly pure gDNA from Gram-positive and Gram-negative bacteria, in less than 20 min.

FGLamide Allatostatin genes in Arthropoda: introns early or late?

FGLamide allatostatins are invertebrate neuropeptides which inhibit juvenile hormone biosynthesis in Dictyoptera and related orders and also show myomodulatory activity. The FGLamide allatostatin (AST) gene structure in Dictyoptera is intronless within the ORF, whereas in 9 species of Diptera, the FGLamide AST ORF has one intron. To investigate the evolutionary history of AST intron structure, (intron early versus intron late hypothesis), all available Arthropoda FGLamide AST gene sequences were examined from genome databases with reference to intron presence and position/phase. Three types of FGLamide AST ORF organization were found: intronless in I. scapularis and P. humanus corporis; one intron in D. pulex, A. pisum, A. mellifera and five Drosophila sp.; two introns in N. vitripennis, B. mori strains, A. aegypti, A. gambiae and C. quinquefasciatus. The literature suggests that for the majority of genes examined, most introns exist between codons (phase 0) which may reflect an ancient function of introns to separate protein modules. 60% of the FGLamide AST ORFs introns were between the first and second base within a codon (phase 1), 28% were between the second and third nucleotides within a codon (phase two) and 12% were phase 0. As would be required for correct intron splicing consensus sequence, 84% of introns were in codons starting with guanine. The positioning of introns was a maximum of 9 codons from a dibasic cleavage site. Our results suggest that the introns in the analyzed species support the intron late model.

Presence, processing, and localization of mouse ADAM15 during sperm maturation and the role of its disintegrin domain during sperm-egg binding.

Successful fertilization requires gametes to complete several stages, beginning with maturation and transport along the male and female reproductive tracts and ending with the interaction between the sperm and the egg. This last step involves sperm-egg adhesion and membrane fusion. ADAMs (disintegrin and metalloprotease domain proteins) are a family of membrane-anchored glycoproteins that are thought to play diverse roles in cell-cell adhesion through their interaction with integrins. This study analyzes the presence, location, processing, and possible role of ADAM15 in mouse sperm. The presence of ADAM15 in mouse spermatozoa was detected by Western blotting, which revealed that ADAM15 is post-translationally processed, during epididymal sperm maturation and the acrosome reaction. The 35 kDa antigen present in the acrosome-reacted sperm is the last proteolytic product of the 110/75 kDa ADAM15 found in non-capacitated sperm. This 35 kDa protein contains the disintegrin domain. By indirect immunofluorescence, ADAM15 was identified in the acrosomal region and along the flagellum of mouse spermatozoa. In acrosome-reacted sperm, ADAM15 was lost from the acrosomal region, but remained diffusely distributed throughout the head and flagellum. Furthermore, the ADAM15 disintegrin domain (RPPTDDCDLPEF) partially inhibited fusion and almost completely inhibited sperm-oolemma adhesion. In conclusion, our data indicate that ADAM15 is present in the testis and in spermatozoa from the caput, corpus, and cauda epididymis, as well as in non-capacitated and acrosome-reacted gametes. Results also indicate that ADAM15 is processed during epididymal maturation and acrosome reaction and that it may play a role during sperm-egg binding.

Loss of DNA: a plausible molecular level explanation for crustacean neuropeptide gene evolution.

Alignment of nucleotides of APGWamide, RPCH and AKH genes gives region stretches (common regions) present in all family member variants. Common regions were separated by gap sections in the larger variants of family members. Consensus sequences for single polynucleotides from virtual hybrid molecules of DNA were obtained by joining the common regions of DNA and deleting the extra DNA nucleotides. Conceptual translation of these virtual hybrids resulted in polypeptides similar to APGWamide, RPCH and the AKH pre-pro-peptide. Virtual polypeptides were also similar to LWamide and RFamide along hydras to mammals. DNA loss probably explains the origin of neuropeptides.

Role of dystrophins and utrophins in platelet adhesion process.

Platelets are crucial at the site of vascular injury, adhering to the sub-endothelial matrix through receptors on their surface, leading to cell activation and aggregation to form a haemostatic plug. Platelets display focal adhesions as well as stress fibres to contract and facilitate expulsion of growth and pro-coagulant factors contained in the granules and to constrict the clot. The interaction of F-actin with different actin-binding proteins determines the properties and composition of the focal adhesions. Recently, we demonstrated the presence of dystrophin-associated protein complex corresponding to short dystrophin isoforms (Dp71d and Dp71) and the uthophin gene family (Up400 and Up71), which promote shape change, adhesion, aggregation, and granule centralisation. To elucidate participation of both complexes during the platelet adhesion process, their potential association with integrin beta-1 fraction and the focal adhesion system (alpha-actinin, vinculin and talin) was evaluated by immunofluorescence and immunoprecipitation assays. It was shown that the short dystrophin-associated protein complex participated in stress fibre assembly and in centralisation of cytoplasmic granules, while the utrophin-associated protein complex assembled and regulated focal adhesions. The simultaneous presence of dystrophin and utrophin complexes indicates complementary structural and signalling mechanisms to the actin network, improving the platelet haemostatic role.

Circadian oscillations of RPCH gene expression in the eyestalk of the crayfish Cherax quadricarinatus.

The RPCH and beta-actin cDNAs from the crayfish Cherax quadricarinatus were amplified, cloned and sequenced. The primary structure sequences of these cDNAs were compared to other members of the AKH/RPCH family. Fluctuations in the amount of the C. quadricarinatus RPCH and beta-actin mRNAs, as cDNAs, were quantified every 3h by RT-PCR. Single cosinor analysis supports the notion of beta-actin and RPCH mRNA circadian behavior in animals subjected to 12h:12h light/dark regimes. In constant darkness RPCH mRNA concentration changes to ultradian cycles.

Platelet adhesion: structural and functional diversity of short dystrophin and utrophins in the formation of dystrophin-associated-protein complexes related to actin dynamics.

Platelets are dynamic cell fragments that modify their shape during activation. Utrophin and dystrophins are minor actin-binding proteins present in muscle and non-muscle cytoskeleton. In the present study, we characterised the pattern of Dp71 isoforms and utrophin gene products by immunoblot in human platelets. Two new dystrophin isoforms were found, Dp71f and Dp71 d, as well as the Up71 isoform and the dystrophin-associated proteins, alpha and beta -dystrobrevins. Distribution of Dp71d/Dp71delta110m, Up400/Up71 and dystrophin-associated proteins in relation to the actin cytoskeleton was evaluated by confocal microscopy in both resting and platelets adhered on glass. Formation of two dystrophin-associated protein complexes (Dp71d/Dp71delta110m approximately DAPC and Up400/Up71 approximately DAPC) was demonstrated by co-immunoprecipitation and their distribution in relation to the actin cytoskeleton was characterised during platelet adhesion. The Dp71d/Dp71delta100m approximately DAPC is maintained mainly at the granulomere and is associated with dynamic structures during activation by adhesion to thrombin-coated surfaces. Participation of both Dp71d/Dp71delta110m approximately DAPC and Up400/Up71 approximately DAPC in the biological roles of the platelets is discussed.

A possible molecular ancestor for mollusk APGWamide, insect adipokinetic hormone, and crustacean red pigment concentrating hormone.

Precursor structures of various members of the neuropeptide family adipokinetic hormone/red pigment concentrating hormone (AKH/RPCH) of mandibular arthropods and the APGWamide family of mollusks were compared. Amino acid alignments showed a common overall architecture (signal peptide, active peptide, related peptide), with a similar alpha helix-random coil secondary structure. DNA sequence alignments revealed close similarities between the genes encoding for the peptides of the two families. The APGWamide genes are larger than the AKH/RPCH genes. The sequence environment occupied by introns is similar in AKH/RPCH and APGWamide genes. Such similarities suggest that these peptide families might have been originated by gene rearrangements from a common ancestor having either an AKH/RPCH/APGWamide-like structure or both an AKH/RPCH-like and an APGWamide-like structures. In the former model, DNA fragments could have been gained when the ancestor evolved to mollusks and it could have lost nucleotides when the progression to mandibular arthropods took place. In the second model, AKH/RPCH-like structures could have been fused during evolution toward mandibular arthropods, whereas in mollusks they could have been lost with the possible amplification of the APGWamide-like structure. Loss of domains in exon 1 may have originated the signal peptide and the first codon of the active RPCH. In exon 2, loss of domains possibly determined the junctions of codons 2 to 5 with the loss of a APGWamide copy; exon 3 underwent fewer variations. The similarity of the mollusk APGWamide precursors is closer to that of the RPCH family than the insect AKH family, indicating an earlier evolutionary departure.