In silico analysis of regulatory and structural motifs of the ovine HSP90AA1 gene

Gene promoters are essential regions of DNA where the transcriptional molecular machinery to produce RNA molecules is recruited. In this process, DNA epigenetic modifications can acquire a fundamental role in the regulation of gene expression. Recently, in a previous work of our group, functional features and DNA methylation involved in the ovine HSP90AA1 gene expression regulation have been observed.Publishe

Ovine HSP90AA1 Expression Rate Is Affected by Several SNPs at the Promoter under Both Basal and Heat Stress Conditions

The aim of this work was to investigate the association between polymorphisms located at the HSP90AA1 ovine gene promoter and gene expression rate under different environmental conditions, using a mixed model approach. Blood samples from 120 unrelated rams of the Manchega sheep breed were collected at three time points differing in environmental conditions. Rams were selected on the basis of their genotype for the transversion G/C located 660 base pairs upstream the gene transcription initiation site. Animals were also genotyped for another set of 6 SNPs located at the gene promoter. Two SNPs, G/C−660 and A/G−444, were associated with gene overexpression resulting from heat stress. The composed genotype CC−660-AG−444 was the genotype having the highest expression rates with fold changes ranging from 2.2 to 3.0. The genotype AG−522 showed the highest expression levels under control conditions with a fold change of 1.4. Under these conditions, the composed genotype CC−601-TT−524-AG−522-TT−468 is expected to be correlated with higher basal expression of the gene according to genotype frequencies and linkage disequilibrium values. Some putative transcription factors were predicted for binding sites where the SNPs considered are located. Since the expression rate of the gene under alternative environmental conditions seems to depend on the composed genotype of several SNPs located at its promoter, a cooperative regulation of the transcription of the HSP90AA1 gene could be hypothesized. Nevertheless epigenetic regulation mechanisms cannot be discarded.Publishe

Ovine HSP90AA1 gene promoter: functional study and epigenetic modifications

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Everything matters in a promoter. New insights on the ovine HSP90AA1gene regulation: abstract

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Differences in the Ovine HSP90AA1 Gene Expression Rates Caused by Two Linked Polymorphisms at Its Promoter Affect Rams Sperm DNA Fragmentation under Environmental Heat Stress Conditions

Heat shock (HS) is one of the best-studied exogenous cellular stresses. Almost all tissues, cell types, metabolic pathways and biochemical reactions are affected in greater or lesser extent by HS. However, there are some especially thermo sensible cellular types such as the mammalian male germ cells. The present study examined the role of three INDELs in conjunction with the -660G/C polymorphism located at the HSP90AA1 promoter region over the gene expression rate under HS. Specially, the -668insC INDEL, which is very close to the -660G/C transversion, is a good candidate to be implied in the transcriptional regulation of the gene by itself or in a cooperative way with this SNP. Animals carrying the genotype II-668 showed higher transcription rates than those with ID-668 (FC = 3.07) and DD-668 (FC = 3.40) genotypes for samples collected under HS. A linkage between gene expression and sperm DNA fragmentation was also found. When HS conditions were present along or in some stages of the spermatogenesis, alternative genotypes of the -668insC and -660G/C mutations are involved in the effect of HS over sperm DNA fragmentation. Thus, unfavorable genotypes in terms of gene expression induction (ID-668GC-660 and DD-668GG-660) do not produce enough mRNA (stored as messenger ribonucleoprotein particles) and Hsp90α protein to cope with future thermal stress which might occur in posterior stages when transcriptional activity is reduced and cell types and molecular processes are more sensible to heat (spermatocytes in pachytene and spermatids protamination). This would result in the impairment of DNA packaging and the consequent commitment of the events occurring shortly after fertilization and during embryonic development. In the short-term, the assessment of the relationship between sperm DNA fragmentation sensitivity and ram’s fertility will be of interest to a better understanding of the mechanisms of response to HS and its consequences on animal production and reproduction performance.Publishe

Influence of the Temperature and the Genotype of the HSP90AA1 Gene over Sperm Chromatin Stability in Manchega Rams

The present study addresses the effect of heat stress on males' reproduction ability. For that, we have evaluated the sperm DNA fragmentation (DFI) by SCSA of ejaculates incubated at 37°C during 0, 24 and 48 hours after its collection, as a way to mimic the temperature circumstances to which spermatozoa will be subject to in the ewe uterus. The effects of temperature and temperature-humidity index (THI) from day 60 prior collection to the date of semen collection on DFI were examined. To better understand the causes determining the sensitivity of spermatozoa to heat, this study was conducted in 60 males with alternative genotypes for the SNP G/C−660 of the HSP90AA1 promoter, which encode for the Hsp90α protein. The Hsp90α protein predominates in the brain and testis, and its role in spermatogenesis has been described in several species. Ridge regression analyses showed that days 29 to 35 and 7 to 14 before sperm collection (bsc) were the most critical regarding the effect of heat stress over DFI values. Mixed model analyses revealed that DFI increases over a threshold of 30°C for maximum temperature and 22 for THI at days 29 to 35 and 7 to 14 bsc only in animals carrying the GG−660 genotype. The period 29–35 bsc coincide with the meiosis I process for which the effect of the Hsp90α has been described in mice. The period 7–14 bsc may correspond with later stages of the meiosis II and early stages of epididymal maturation in which the replacement of histones by protamines occurs. Because of GG−660 genotype has been associated to lower levels of HSP90AA1 expression, suboptimal amounts of HSP90AA1 mRNA in GG−660 animals under heat stress conditions make spermatozoa DNA more susceptible to be fragmented. Thus, selecting against the GG−660 genotype could decrease the DNA fragmentation and spermatozoa thermal susceptibility in the heat season, and its putative subsequent fertility gainsPublishe

The European Reference Genome Atlas: piloting a decentralised approach to equitable biodiversity genomics.

ABSTRACT: A global genome database of all of Earth’s species diversity could be a treasure trove of scientific discoveries. However, regardless of the major advances in genome sequencing technologies, only a tiny fraction of species have genomic information available. To contribute to a more complete planetary genomic database, scientists and institutions across the world have united under the Earth BioGenome Project (EBP), which plans to sequence and assemble high-quality reference genomes for all ∼1.5 million recognized eukaryotic species through a stepwise phased approach. As the initiative transitions into Phase II, where 150,000 species are to be sequenced in just four years, worldwide participation in the project will be fundamental to success. As the European node of the EBP, the European Reference Genome Atlas (ERGA) seeks to implement a new decentralised, accessible, equitable and inclusive model for producing high-quality reference genomes, which will inform EBP as it scales. To embark on this mission, ERGA launched a Pilot Project to establish a network across Europe to develop and test the first infrastructure of its kind for the coordinated and distributed reference genome production on 98 European eukaryotic species from sample providers across 33 European countries. Here we outline the process and challenges faced during the development of a pilot infrastructure for the production of reference genome resources, and explore the effectiveness of this approach in terms of high-quality reference genome production, considering also equity and inclusion. The outcomes and lessons learned during this pilot provide a solid foundation for ERGA while offering key learnings to other transnational and national genomic resource projects.info:eu-repo/semantics/publishedVersio

Estudio de la función y regulación del gen ovino "HSP90AA1"

El cambio climático, el aumento de la temperatura global y la desertificación de nuestro planeta hacen necesaria la búsqueda de especies ganaderas rústicas y adaptadas a estas nuevas condiciones ambientales, y cuya capacidad productiva no se vea mermada por dichos factores. La especie ovina fue una de las primeras especies domesticadas. Este pequeño rumiante está muy bien adaptado a climas agrestes y áridos y generalmente se explota por su triple aptitud productiva de lana, leche y carne. Todos los organismos vivos poseen una serie de mecanismos para afrontar los cambios de temperatura que se producen en el medio ambiente en que se desarrollan denominados respuesta al estrés térmico (HSR). La HSR activa una serie de genes, entre los cuales tiene gran importancia aquellos que codifican las proteínas de shock térmico (HSP). Las HSPs se encargan de restaurar la homeostasis celular, alterada como consecuencia del estrés por calor (HS). En concreto, la hsp90α realiza esta función identificando proteínas cuya conformación se ve alterada por el aumento de la temperatura para su posterior reparación o destrucción. El gen HSP90AA1 que codifica la Hsp90α, ha sido caracterizado y mapeado en la especie ovina. El presente estudio, se ha focalizado en la caracterización de la región promotora de este gen que es altamente polimórfica. Con el fin de analizar el posible efecto de 11 polimorfismos sobre la regulación del gen, se han realizado estudios de expresión génica con muestras biológicas recogidas en momentos de distintas condiciones ambientales. Se ha podido constatar que el genotipo CC de la transversión g.660G>C incrementa la expresión del gen tanto en condiciones de termo neutralidad como de HS. Además, que la presencia en homocigosis de la inserción de una citosina, g.667_668insC, incrementa hasta tres veces la tasa de transcripción del gen solo bajo condiciones de estrés térmico. El promotor del gen HSP90AA1 ha sido caracterizado como un promotor híbrido que no solo contiene caja TATA, sino que además posee una isla CpG reguladora. Este hecho confiere a este tipo de promotores una mayor plasticidad respecto a la regulación de su transcripción. Además, dentro de esta isla CpG es donde se encuentran los polimorfismos detectados, constatándose que el SNP g.660G>C genera un sitio de metilación de novo especifico del alelo G. Mediante experimentos in vitro se pudo comprobar que la combinación de dos citosinas, g.667_668insC y C-660, producen la mayor tasa de transcripción del gen bajo HS. Se ha determinado la presencia de los 11 polimorfismos detectados en 31 razas ovinas constatándose la existencia de una correlación significativa entre las frecuencias de aquellos ligados a la respuesta a estrés térmico y las variables climáticas preponderantes de los lugares de origen de algunas de las razas ovinas estudiadas. La comparación de la secuencia del promotor del gen HSP90AA1 en 9 especies domesticas y salvajes de la subfamilia Caprinae ha permitido comprobar la presencia de varios de los polimorfismos en dichas especies y obtener resultados sorprendentes en cuanto a sus posibles relaciones filogenéticas. Finalmente, y como resultado más directamente aplicable al sector de producción ovina, se han correlacionado los genotipos de los polimorfismos ligados a la HSR con la tasa de fragmentación del ADN espermático de los carneros medida en momentos de diferentes condiciones climáticas. Se ha observado que existen fases en el proceso de la espermatogénesis, en las que la temperatura ambiental es un factor crítico en el óptimo desarrollo de las células germinales masculinas. Estos resultados, indican que la gran variabilidad polimórfica presente en la región promotora del gen HSP90AA1 tiene una gran implicación a nivel molecular con consecuencias sobre la variabilidad de un carácter reproductivo