Identification of endocytosis-associated genes, and full-length transcripts, in Apis mellifera ligustica
Author of the article:FAN Xiao-Xue, WANG Si-Yi, ZHU Le-Ran, JING Xin, GUO Si-Jia, NIU Qing-Sheng, JIANG Hai-Bin, XU Xi-Jia
Author's Workplace：College of Animal Sciences (College of Bee Science), Fujian Agriculture and Forestry University, Fuzhou 350002, China; Apitherapy Research Institute, Fujian Agriculture and Forestry University, Fuzhou 350002, China
Key Words：Apis mellifera ligustica; endocytosis; Nanopore sequencing; full-length transcript; alternative splicing
Abstract：
[Objectives] To identify, analyze and confirm the genes and full-length transcripts associated with endocytosis in Apis mellifera ligustica using previously obtained Nanopore long-read sequencing data, thereby providing a foundation for further functional study of these genes. [Methods] All full-length transcripts available for Apis mellifera ligustica were Blast-searched in the Nr and KEGG databases to screen out endocytosis-related genes and full-length transcripts. Structures of annotated genes were optimized by using gffcompare software to compare endocytosis-associated full-length transcripts with annotated transcripts in the A. mellifera reference genome (Amel_HAv3.1). Types of AS events in genes related to endocytosis were identified using Astalavista software. Visualization of isoforms’ structure was performed with an IGV browser. The authenticity of AS events was verified using PCR. Prediction and investigation of APA sites were done using a TAPIS pipeline, followed by identification of motifs upstream of APA sites with MEME software. [Results] In total, 170 genes and 991 full-length transcripts relevant to endocytosis were identified. Structures of 89 annotated genes were optimized, among these, 5' ends of 36 genes were prolonged, 3' ends of 35 genes were prolonged, and both 5' ends and 3' ends of 15 genes were prolonged. In addition, 666 AS events in 28 genes were identified, the most common of which was an alternative 3' splice site. PCR results validated the authenticity of four randomly selected AS events involving an integrin beta-nu-like isoform, ras-like GTP-binding protein and Ras-like GTP-binding protein. Moreover, 120 genes were identified containing one or more APA sites in endocytosis, the most abundant of which were genes containing three APA sites (14, 11.67%). Several motifs upstream of APA sites were observed, with the consensus sequence: NBMNHHHBMNSNNCBNVSNNNNNNNNNNNVNNNN. [Conclusion] 170 genes and 991 full-length transcripts associated with endocytosis in A. m. ligustica were identified. A number of AS events in endocytosis-related genes that also contained abundant APA sites were detected, and structures of endocytosis- related genes annotated in the reference genome of A. mellifera were optimized.