Project description:The trypanosomatid protozoan parasite Leishmania has a significant impact on human health globally. Understanding the pathways associated with virulence within this significant pathogen is critical for identifying novel vaccination and chemotherapy targets. Within this study we leverage an ultradeep proteomic approach to improve our understanding of two virulence associated genes in Leishmania; the Golgi Mannose/Arabinopyranose/Fucose nucleotide-sugar transporter LPG2, and the mitochondrial fucosyltransferase FUT1. Using deep peptide fractionation followed by complementary fragmentation approaches with higher energy collisional dissociation (HCD) and Electron-transfer dissociation (ETD) allowed the identification of over 6500 proteins, nearly doubling the experimentally observed Leishmania major proteome. This deep proteomic analysis revealed significant quantitative differences in both lpg2- and fut1s mutants with FUT1-dependent changes linked to marked alterations within mitochondrial associated proteins while LPG2-dependent changes impacted multiple aspects of the secretory pathway. While FUT1 has been shown to fucosylate peptides in vitro, no evidence for protein fucosylation was identified within our ultradeep analysis nor did we observe fucosylated glycans within Leishmania glycopeptides isolated using HILIC enrichment. Combined this work provides a critical proteomic resource for the community on the observable Leishmania proteome as well as highlights phenotypic changes associated with LPG2/FUT1 which may guide the development of future therapeutics.
Project description:This data was generated to identify the molecular pathways responsible for nitrous oxide synthesis by the green algae Chlamydomonas reinhardtii, when supplied with nitrite under aerobic conditions (oxia). RNA samples were collected at three time points, 15 min, 3 hours, and 24 hours after the start of the experiment. The control and treatment groups were grown under the same conditions, except treatment group was supplied with 10mM nitrite at time 0. Illumina TruSeq stranded RNA libraries were synthesised from the resulting RNA before sequencing on a HiSeq2500 (125bp). The resulting sequence run generated 241,151,809 paired-end 125bp reads, of which 200,946,839 remained following quality filtering. The short data was mapped to the published genome and read counts were generated with HT-Seq count with the default settings. The raw read count data was analysed by DESeq2 in order to identify genes differentially expressed during nitrous oxide production.