Identification of Sheep Hair Follicles Development-related lncRNAs Based on RNA-seq Technology

doi:10.7668/hbnxb.20194552

Abstract

Abstract:

The hair follicles of Dorper sheep are characterised by cyclic growth and development,which is generally divided into three periods:anagen,catagen,and telogen.This study aims to explore the regulatory mechanism of key lncRNAs as ceRNA associated with hair follicles anagen and telogen as screened by previous studies.Ten Dorper ewes (shedding group,S group) with extreme shedding phenotypes and 10 non-shedding ewes (non-shedding group,N group) were selected under the same feeding conditions,with similar body weights and sizes,and about 2 years old,and the best 5 shedding ewes and 3 non-shedding ewes were selected for transcriptome sequencing after phenotypic observation.The ceRNA regulatory network of key lncRNAs was constructed by bioinformatics analysis methods such as target prediction,expression pattern analysis,and GO and KEGG enrichment analysis,to further investigate the regulatory mechanisms of key lncRNAs.The results showed a total of 262 mRNAs were screened by lncRNA-miRNA and miRNA-mRNA target prediction analysis,of which 135 mRNAs were consistent with the expression patterns of the corresponding lncRNAs (19 A pattern (Anagen,highly expressed in anagen) and 116 T pattern (Telogen,highly expressed in telogen)).GO and KEGG enrichment analysis of these 135 mRNAs revealed that some genes associated with development of hair follicles,such as CTNNB1,FGF22,FGF5,FZD3,JAK3,Lpar6,NGFR,PAK1,EIF4E,and Wnt10b,were enriched in Rap1,MAPK,Wnt,PI3K-Akt,Ras,mTOR,Jak-STAT,Hippo and other signaling pathways,which were associated with development of hair follicles.A ceRNA regulatory network consisting of 10 lncRNAs,11 miRNAs and 10 mRNAs was finally constructed.The qRT-PCR of several randomly selected differentially expressed lncRNAs and differentially expressed genes showed that the expression trend was basically consistent with that of the transcriptome sequencing results,indicating that the sequencing result was reliable.These 10 lncRNAs obtained from the study together with their target-regulated miRNAs and mRNAs can be the focus of research on the development of hair follicles in sheep.

Key words: Sheep, lncRNA, ceRNA, Hair follicle, Anagen, Telogen

SUN Haoran, LI Xinhai. Identification of Sheep Hair Follicles Development-related lncRNAs Based on RNA-seq Technology[J]. Acta Agriculturae Boreali-Sinica, 2024, 39(1): 211-218. doi: 10.7668/hbnxb.20194552.

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http://www.hbnxb.net/EN/Y2024/V39/I1/211

Figures/Tables 6

Tab.1 Information of fluorescent quantitative primers

类型 Type	基因/lncRNAs Genes/lncRNAs	引物序列(5'—3') Primer sequence (5'—3')	引物长度/bp Primer length
内参基因	GAPDH	F: TCCACGGCACAGTCAAGG	239
Internal reference gene		R: CACGCCCATCACAAACAT
LncRNAs	MSTRG.12818.1	F: GGACGTGCCTTTAGAATGATGA	228
		R: TACCTGTGTTTGTGTCGCTGTG
	XR_003588784.1	F: TCCCGTCTTCCTGTTCGCCCTCTT	130
		R: TCTCGCCTTGCCCGCTCTCACCCT
基因	GPX4	F: CAGGAGCCAGGGAGTAATGCGGAGAT	208
Genes		R: CCACACAGCCGTTCTTATCAATCAGG
	FABP4	F: AAGAAGTGGGTGTGGGCTTTGCTA	244
		R: GTTGATTTCCCATCCCAGTTTTGT
	KRT17	F: CACCATCCGCCACTTCTCC	273
		R: GCTTTCTCGCCTCCCACCA
	KRT27	F: TCTCTCTGGCAATGAGAAGGTGACCA	186
		R: ACCGTGAAGTATCTGCTATAATCGTG

Fig.1 Expression patterns of target genes conforming to the A or T pattern A.The 19 target genes conforming to the A pattern (Anagen,highly expressed in anagen);B.The 116 target genes conforming to the T pattern (Telogen,highly expressed in telogen).

Fig.2 Secondary classification histogram and for GO enrichment analysis of 135 target genes conforming to A or T pattern

Fig.3 Significance bubble plot for KEGG enrichment analysis of 135 target genes that fit the A or T pattern 1.Human papillomavirus infection;2.Kaposi sarcoma-associated herpesvirus infection;3.Proteoglycans in cancer;4.Human cytomegalovirus infection;5.Human T-cell leukemia virus 1 infection;6.Pathways in cancer;7.PI3K-Akt signaling pathway;8.Wnt signaling pathway;9.Hepatitis B;10.Phospholipase D signaling pathway;11.Chemokine signaling pathway;12.Neurotrophin signaling pathway;13.Th1 and Th2 cell differentiation;14.Th17 cell differentiation;15.PD-L1 expression and PD-1 checkpoint pathway in cancer;16.MAPK signaling pathway;17.Melanogenesis;18.Ras signaling pathway;19.Rap1 signaling pathway;20.NF-kappa B signaling pathway.

Fig.4 Diagram of the ceRNA regulatory network of crucial lncRNAs Green circles.Target genes;Red diamonds.miRNAs;Yellow triangles.lncRNAs.

Fig.5 Comparison between qRT-PCR and RNA-seq results

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