Molecular Mechanism Of Porcupine Quills Growth Based On Transcriptome Data

Chinese subspecies of Malayan porcupine(Hystrix brachyura subcristata)has a complex coat type,and the back spines formed by the specialization of body hair are one of its more obvious features,which are essentially the product of hair follicle cycle growth and the result of coat variation,but the formation mechanism has not been clarified.There are about 30,000 spines on each porcupine.Comparative studies on skin,hair and spines in different parts of porcupine are helpful to understand the relevant regulatory factors and mechanisms involved in hair formation.In this study,3 porcupine calves were selected within 3 days of birth,and transcriptome sequencing was performed on samples of back skin,belly skin,back spines and belly hair.Three sets of replicates were designed for each group of samples to analyze the differentially expressed genes affecting hair formation at different parts.GO functional annotation and KEGG signaling pathway analysis were performed on differentially expressed genes between sample groups.Preliminary screening of regulatory genes involved in different hair formation processes.The main results are as follows:(1)Transcriptome sequencing is performed on the Dorsal skin samples,abdominal skin samples Ventral,dorsal Quill samples and abdominal Hair samples using the dorsal high throughput sequencing platform.There were an average of 42563956.67 clean reads in the spiny back sample group,42262039.33 clean reads in the back skin sample group and 42859813.33 clean reads in the abdominal skin sample group.Due to insufficient amount of abdominal hair extraction,only 43483500 clean reads were obtained from one sample,and subsequent analysis was based on clean Reads.After assembling and splicing the samples,348,852 Trinity pieces and 197,738 Unigene pieces were obtained.After assembly,the maximum length is 40,932 bp and 40,932 bp respectively,the minimum length is 201 bp,and the average length is 1,409.82 bp and 843.44 bp respectively.A total of197738 comments were obtained,27807 to the GO database and 15184 to the KEGG database.(2)Through the enrichment of differentially expressed genes between the Quill sample group and Dorsal sample group,13,670 differentially expressed genes were screened,among which 5099 up-regulated genes and 8571 down-regulated genes.Through the enrichment of differentially expressed genes between the Quill3 vs Hair samples,19,212 differentially expressed genes were screened,11663 were up-regulated and 7549 were down-regulated.Enrichment of differential genes between Dorsalvs Ventral samples.1426 differentially expressed genes were screened,with 1080 up-regulated genes and 346down-regulated differentially expressed genes.Through the enrichment of differentially expressed genes between the Quillvs Ventral samples,15974 differentially expressed genes were screened,among which 7644 were up-regulated and 8330 were down-regulated.(3)The differential expression genes dorsal in the Quillvs Dorsal sample group were enriched to 57 entries in the GO database and annotated to 102 entries in the KEGG database.Fifty-seven differential gene enrichment notes of Quill3 vs Hair3 sample group were recorded in the GO database and 31 in the KEGG database.46 notes of differential gene enrichment in the Dorsalvs Ventral sample group were recorded in the GO database and 14 in the KEGG database.Fifty-nine differential gene enrichment notes of Quillvs Ventral sample group were recorded in the GO database and 87 in the KEGG database.(4)198212 differentially expressed genes from dorsal spines and abdominal hair were annotated in NT,NR,BLASTX,Uniprot,PFAM,egg NOG,GO and KEGG databases,and compared with dorsal spines and dorsal skin,and dorsal spines and abdominal skin B3A4,SOX21,CELR1,SHH,TGFB2,SOX18,K1C27,NOTC1,K1C25,K1C27,FGF10,K2C71,and SOX18 and FGF10 were only expressed in dorsal spines.Keratin-related differential genes: KRT26,KRT27,KRT71,KRT72,KRT74,KRT76;Differential genes related to keratin-related proteins were KR241,KR261,KR107,KRA97,KR412,KR122,and transcription factors CUX1,SOX10,LHX9,RUNX3,which were related to hair follicle development,morphogenesis,and differentiation.(5)The differential genes enriched in the GO and KEGG pathways were analyzed.It is mainly enriched by Cell adhesion molecules(CAMs),Pathways in cancer,and AGE-RAGE signaling pathway in diabetic complications diabetic complications,PI3K-Akt signaling pathway,and Wnt signaling pathway.Combined with the reported literature,significant differential genes in each pathway were analyzed,and it was found that Wnt2,FGF5,Wnt5,DKK1,NFATC1,EGF,EGFR,TGF-β,EDA and other genes involved in hair follicle differentiation may be related to the formation of porcupine spines.