RNA-seq of mutant Mycobacterium neoaurum producing steroid intermediates compared with wild-type strain
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina Hiseq 2000, Illumina HiSeq 2000
ORGANISM(S): Mycobacterium neoaurum
SUBMITTER: Min Liu
PROVIDER: E-MTAB-6789 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Background: In recent years, an innovative strategy using laser microdissection and mass spectrometry markedly expanded the landscape of antigens associated with membranous nephropathy (MN). Specific associations with phenotypes, diseases and sometimes reversible triggers led to a novel antigen-based classification of MN, paving the way for precision medicine and stressing the need for more routine use of proteomics in MN. Methods: To explore the proteomic landscape of human glomeruli and identify podocyte antigens and disease mechanisms in MN, we expanded the original technique to an integrative approach combining laser capture microdissection, next-generation mass spectrometry and computational analysis. Next to conventional data-dependent acquisition (DDA), we used and assessed the diagnostic yield of the more comprehensive data-independent acquisition (DIA) mass spectrometry, which enables the detection and quantification of every peptide in a sample irrespective of its level of abundance or m/z value. Our proteomic pipeline was applied to residual material from kidney biopsies in 64 individuals, including 31 healthy controls; 5 disease controls; 5 PLA2R-associated MN; and 23 PLA2R-negative MN. Results: Unbiased analyses confirmed the significant enrichment in PLA2R, IgG4 and complement proteins in glomeruli from patients with PLA2R-MN compared with healthy and disease controls, while molecular characterization of complement fragments provided evidence for complement activation in PLA2R-MN. Compared to DDA, DIA mass spectrometry increased the number of glomerular proteins (~3800 vs. ~1200) identified in healthy glomeruli; allowed the detection all known antigens except NELL1 in normal glomeruli; and increased the detection rate of podocyte antigens from 50% to >80% in PLA2R-negative MN. Conclusions: This proof-of-concept study suggests that an integrative approach combining laser microdissection, DIA mass spectrometry and computational biology is a powerful tool, with translational potential, to identify podocyte antigens and unravel disease mechanisms in MN.
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Project description:Iron and manganese are part of a small group of transition metals required for photosynthetic electron transport. Here, we present evidence for a functional link between iron and manganese homeostasis. In the unicellular cyanobacterium, Synechocystis sp. PCC 6803 Fe and Mn deprivation resulted in distinct modifications of the function of the photosynthetic apparatus. For example, iron limitation modifies the rate of QA re-oxidation in photosystem II, a complex that contains more Mn than Fe. The intracellular elemental quotas of Fe and Mn are also linked. Fe limitation reduces the intracellular Mn quota. Mn limitation did not exert a reciprocal effect on Fe quotas. Microarray analysis comparing Mn and Fe limitation revealed a stark difference in the extent of the transcriptional response to the two limiting conditions, reflective of the physiological data. The effects of Fe limitation on the transcriptional network are widespread while the effects on Mn limitation are highly specific. Our analysis also revealed an overlap in the transcriptional response of specific Fe and Mn transporters. This overlap provides a framework for explaining Fe limitation induced changes in Mn quotas. Fe transporters can serve as a low affinity Mn transport system. Under iron limitation the specificity of the Fe transport system changes, making it a less efficient Mn transport system. We monitored the gene expression of Synechocystis PCC6083 at standard conditions and after 2 days of iron limitation (0Fe), manganese limitation (0Mn) and combined iron and manganese limitation (0Fe0Mn). Each timepoint and condition was sampled in triplicates. Due to strong deviations in one of the three repeats for the 0Mn and 0Fe0Mn conditions, the corresponding replicates were excluded from further analysis.
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Project description:Background: In recent years, an innovative strategy using laser microdissection and mass spectrometry markedly expanded the landscape of antigens associated with membranous nephropathy (MN). Specific associations with phenotypes, diseases and sometimes reversible triggers led to a novel antigen-based classification of MN, paving the way for precision medicine and stressing the need for more routine use of proteomics in MN. Methods: To explore the proteomic landscape of human glomeruli and identify podocyte antigens and disease mechanisms in MN, we expanded the original technique to an integrative approach combining laser capture microdissection, next-generation mass spectrometry and computational analysis. Next to conventional data-dependent acquisition (DDA), we used and assessed the diagnostic yield of the more comprehensive data-independent acquisition (DIA) mass spectrometry, which enables the detection and quantification of every peptide in a sample irrespective of its level of abundance or m/z value. Our proteomic pipeline was applied to residual material from kidney biopsies in 64 individuals, including 31 healthy controls; 5 disease controls; 5 PLA2R-associated MN; and 23 PLA2R-negative MN. Results: Unbiased analyses confirmed the significant enrichment in PLA2R, IgG4 and complement proteins in glomeruli from patients with PLA2R-MN compared with healthy and disease controls, while molecular characterization of complement fragments provided evidence for complement activation in PLA2R-MN. Compared to DDA, DIA mass spectrometry increased the number of glomerular proteins (~3800 vs. ~1200) identified in healthy glomeruli; allowed the detection all known antigens except NELL1 in normal glomeruli; and increased the detection rate of podocyte antigens from 50% to >80% in PLA2R-negative MN. Conclusions: This proof-of-concept study suggests that an integrative approach combining laser microdissection, DIA mass spectrometry and computational biology is a powerful tool, with translational potential, to identify podocyte antigens and unravel disease mechanisms in MN.
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