Essay On Extracellular Trafficking

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Intracellular trafficking is a complex and dynamic process involving the participation of many proteins. Multiple domains of Cin1 allow its interactions with proteins involved in endocytosis and exocytosis, actin cytoskeleton maintenance, and signaling events. In Cn, virulence factors including melanin, the capsule, and proteinases are actively transported via exocytosis. Disruptions of the genes encoding Sav1, Sec6, and Sec14 attenuated the production of capsule, melanin, and phospholipase, respectively {Yoneda, 2006 #5408; Panepinto, 2009 #5385; Chayakulkeeree, 2011 #5686}. In contrast, the disruption of the CIN1 gene encoding Cin1 resulted in not only the defect in cellular transport but also many other defects {Shen, 2010}. How Cin1…show more content…
Approximately 100 ng cDNA each was subject to illumina deep sequencing (BGI, Shenzhen, China) that yieled high quality data. In this aim, we will complete annotation, identification, validation, and functional analysis of all RNA-Seq data.

3a. sRNA analysis To date, exRNA has not been used to investigate intracellular trafficking or pathogenesis in fungi. We first isolated EVs from the cin1 mutant and WT strains and extracted exRNA with Trizol. Since the cin1 mutant is drastically reduced in growth, 3x more culture volume than WT was necessary to obtain a yield comparable to that of WT. Using BGISEQ-500 technology, a total of 27,084,174 cleans reads was obtained from the cin1 mutant while 27,402,307 clean sRNA reads were obtained from WT (JEC21). The length of sRNA is between 18 nt and 30 nt. Initial sRNA annotation provided by BGI suggested that approximately same 96% of sRNA sequence was mapped to available sRNA database including miRBase and Rfam. This and other early reports such as annotation and prediction, miRNA and siRNA target prediction, and validation by expression will be re-analyzed with the help of collaborator Dr. Taylor. Through this effort, we will determine the role of Cin1 in expression and secretion of sRNA.

3b. mRNA and lncRNA analysis In addition to sRNA, mRNA and lncRNA encapsulated in EVs were also included in RNA-Seq. In comparison to mRNA that convey genetic codes from DNA to protein, lncRNA are non-protein
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