بررسی بیوانفورماتیکی تغییرات بیان رونوشت ژن‌های شیردان به آلودگی انگل Heamonchus contortus در گوسفندان مقاوم و حساس

نوع مقاله : مقاله پژوهشی

نویسندگان

1 دانشجوی دکتری، گروه بیوتکنولوژی کشاورزی، پردیس دانشگاهی، دانشگاه گیلان

2 استاد، گروه بیوتکنولوژی کشاورزی، دانشکده کشاورزی، دانشگاه گیلان

3 استادیار، گروه بیوتکنولوژی کشاورزی، دانشکده کشاورزی، دانشگاه گیلان

4 استاد، گروه علوم دامی، دانشکده کشاورزی، دانشگاه گیلان

5 دانشیار، گروه انگل شناسی و قارچ شناسی پزشکی، دانشکده پزشکی، دانشگاه علوم پزشکی گیلان

چکیده

هدف از این مطالعه، انجام یک بررسی سیستماتیک و متاآنالیز داده­های جمع آوری شده از گوسفندان آلوده شده با Heamonchus contortus به عنوان یکی از مهمترین و بیماری­زاترین انگل­های دستگاه گوارش گوسفند و تجزیه و تحلیل کلی تغییر در رونوشت ژن­های شیردان در پاسخ به این عفونت با استفاده از فناوری RNA-seq و ابزارهای بیوانفورماتیک بود. به منظور شناسایی ژن‌ها و مسیرهای مرتبط با عفونت، متاآنالیز با ترکیب مجموعه داده­های مختلف عفونت Heamonchus contortus گوسفند با استفاده از بسته­ رنک پراد نرم افزار R انجام شد و نتایج به­دست آمده برای استحصال اطلاعات بیشتر مورد تجزیه و تحلیل­های پائین­دستی قرار گرفتند. نتایج حاصل از متاآنالیز نشان داد که در مجموع، 1388 ژن بین نژادهای مقاوم و حساس دارای بیان افتراقی (DEGها) بودند. مطابق با نتایج حاصل از تجزیه و تحلیل هستی­شناسی ژن و دانشنامه­ کیوتوی ژن­ها و ژنوم­ها (KEGG)، این ژن­های دارای بیان افتراقی (DEGها) در فرآیندهای زیستی متعددی مانند سوخت و ساز واحدهای یک کربنی، ترجمه، مسیر سیگنالینگ رسپتور سطح سلولی، پاسخ ایمنی، مسیرهای متابولیکی و ... درگیر بودند. با استفاده از تجزیه و تحلیل برهمکنش پروتئین-پروتئین، ژن­های هاب متعددی مانند آلبومین و CD4 شناسایی شدند که ممکن است نشان دهد بهبود پاسخ ایمنی ناشی از افزایش بیان ژن­های مختلف در ایجاد مقاومت تأثیر می­گذارد. نتایج این مطالعه، بینشی کلی از تغییرات در ترانسکریپتوم نژادهای مقاوم و حساس گوسفند و ساز و کارهای مولکولی مقاومت میزبان در نتیجه عفونت Heamonchus contortus ارائه می­دهد که می­تواند مبنایی برای تحقیقات بیشتر در مورد این موضوع فراهم نماید.

کلیدواژه‌ها

موضوعات

عنوان مقاله [English]

Bioinformatics investigation of expression changes of abomasal gene transcripts to Heamonchus contortus infection in resistant and sensitive sheep

نویسندگان [English]

  • R. Seighalani 1
  • H. Samizadeh Lahiji 2
  • F. Rafeie 3
  • M. Mottaghitalab 4
  • M. Galin Sharifdini 5
1 Ph.D. Student, Department of Agricultural Biotechnology, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
2 Professor, Department of Agricultural Biotechnology, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
3 Assistant Professor, Department of Agricultural Biotechnology, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
4 Professor, Department of Animal Science, Faculty of Agricultural Sciences, University of Guilan, Rasht, Iran
5 Associate Professor, Department of Medical Parasitology and Mycology, School of Medicine, Guilan University of Medical Science, Rasht, Iran
چکیده [English]

Introduction: Gastrointestinal nematodes (GIN) represent a major health issue for livestock production systems worldwide. Haemonchus contortus is one of the most pathogenic GIN in small ruminants and causes serious losses to farmers, both in impaired production and in control with anthelmintics. It has long been recognized that differences in host resistance and susceptibility to parasitic infection exist in various sheep breeds and that genetics may play an important role in regulating host resistance, which has spurred efforts to control parasitic infection through selective breeding for naturally resistant sheep. A detailed understanding of the genes and mechanisms involved in expressing a resistant phenotype and the factors that regulate this response would facilitate the identification of candidate genes for selection. Recent advances in high-throughput technology, such as microarrays and RNA sequencing, have made a large number of transcriptome data accessible. As a result, researchers are now able to obtain more reliable results by integrating information from multiple sources. Accordingly, meta-analysis can be used as a useful and powerful tool to identify differential gene expression. It can also find out genes that their products are key molecules in response to the infection and use in animal breeding programs. The purpose of this study was to conduct a systematic review and meta-analysis on data collected from infected sheep with H. contortus and general analysis of changes in abomasal gene transcripts in response to infection using RNA-seq technology and bioinformatics tools.
Materials and methods: In this study, to identify infection-related genes, pathways, and molecular mechanisms underlying host resistance to this parasite, a meta-analysis was performed by combining two different datasets, including 70 samples of sheep H. contortus infection with Rankprod package of R software. After pre-processing, to remove heterogenicity across studies, batch effect correction was performed on gene expression data. The result of the principal component analysis showed that batch correction reduced the batch variation among the datasets. Meta-analysis was carried out and DEGs selected by meta-analysis were further analyzed and characterized. Enrichment analysis, as an efficient method for functional analysis of massive genetic data, was used to determine the biological process, molecular function, and cellular component of DEGs. Moreover, we searched upstream regions of DEGs for over-represented DNA motifs and functional analysis of discovered motifs. To explore the potential interaction network of the DEGs, the protein-protein interaction network among the DEGs was analyzed using the STRING database, which included direct and indirect associations of proteins. After analyzing the result derived from STRING analysis and expression change information for each DEG, the network figure was drawn for the selected DEGs (connected with one or more DEGs) by using the Cytoscape software and hub genes identified with the CytoHubba plugin of Cytoscape.
Results and discussion: Results derived from the meta-analysis showed a total of 1388 differentially expressed genes between resistant and susceptible sheep. Among them, 1137 were significantly upregulated, whereas 251 were downregulated across the datasets. In the identified DEGs, DEGs corresponding to ribosomal protein S3A, lysozyme C-1-like (LOC443320), and heterogeneous nuclear ribonucleoprotein K were the most strongly upregulated ones, while tenascin C and fibromodulin were the most strongly downregulated. Results from enrichment analysis showed these differentially expressed genes (DEGs) were involved in different biological processes such as one-carbon metabolism, translation, cell surface receptor signaling pathway, immune response, metabolic pathways, PPAR signaling pathway, etc. Moreover, searching in upstream regions of DEGs to find DNA motifs, were able to identify eight conserved sequence motifs. The functional analysis of these motifs revealed that they were involved in the positive regulation of gene expression, defense response, positive regulation of immune response, cellular calcium ion homeostasis, etc. Using the protein-protein interaction analysis also identified multiple hub genes such as albumin and CD4 which may show that improved immune response, induced by up-regulation different genes affects the creation of resistance.
Conclusions: The mechanisms of sheep resistance to GIN infections involve complex immune responses. Our results offered overall insight into changes in the transcriptomes of resistant and susceptible sheep and molecular mechanisms of host resistance induced by H. contortus infection. We propose these DEGs as a useful resource of molecular biomarkers and potential candidate genes for breeding programs which can provide a basis for further research on this topic.

کلیدواژه‌ها [English]

  • Differential expression
  • RNA-seq analysis
  • Meta-analysis
  • Gene ontology

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تحقیقات تولیدات دامی
دوره 11، شماره 4
بهمن 1401
صفحه 61-75

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