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Proteome of microparticles derived from Mycobacterium tuberculosis infected macrophages

ABSTRACT: Microparticles (MP) are small membranous particles (100-1000 nm) released under normal steady-state conditions and are thought to provide a communication network between host cells. Previous studies demonstrated that Mycobacterium tuberculosis (M.tb) infection of macrophages increased the release of MPs, and these MPs induced a proinflammatory response from uninfected macrophages in vitro and in vivo following their transfer into uninfected mice. To determine how M.tb infection modulates the protein composition of the MPs, and if this contributes to their proinflammatory properties, we compared the proteomes of MPs derived from M.tb-infected (TBinf-MP) and uninfected human THP-1 monocytic cells. MP proteins were analysed by GeLC-MS/MS with spectral counting revealing 68 proteins with statistically significant differential abundances. The 42 proteins increased in abundance in TBinf-MPs included proteins associated with immune function (7), lysosomal/endosomal maturation (4), vesicular formation (12), nucleosome proteins (4) and antigen processing (9). Prominent among these were the type I interferon inducible proteins, ISG15, IFIT1, IFIT2, and IFIT3. Exposure of uninfected THP-1 cells to TBinf-MPs induced increased gene expression of isg15, ifit1, ifit2, and ifit3 and the release of proinflammatory cytokines. These proteins may regulate the proinflammatory potential of the MPs and provide candidate biomarkers for M.tb infection.

INSTRUMENT(S): LTQ Orbitrap Velos, QSTAR

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Macrophage

DISEASE(S): Tuberculosis

SUBMITTER: Nathan Hare

LAB HEAD: Dr Bernadette Saunders

PROVIDER: PXD002026 | Pride | 2022-02-28

REPOSITORIES: Pride

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			Action	DRS
	20120702gel1.10run1.wiff	Wiff
	20120702gel1.10run1.wiff.scan	Wiff
	20120702gel1.10run2.wiff	Wiff
	20120702gel1.10run2.wiff.scan	Wiff
	20120702gel1.11run1.wiff	Wiff

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Project description:Tissue macrophages (MPs) develop distinctive functions due to unique transcriptional programs driven by local environmental ques. In the intestine, it is known that MPs are heterogeneous cells that differentiate from blood monocytes and embryonic precursors, and are essential for homeostasis with the vast microbiome through their ability to kill invading microbes, clear apoptotic cells, and produce regulatory cytokines. However, the full extent of MP heterogeneity, their developmental relationships, and how the intestinal microbiota affects MP generation is not known. Here, we performed single cell RNA sequencing of colonic myeloid cells from specific pathogen free (SPF) and germ free (GF) C57BL/6 mice and found extensive heterogeneity of colon MP, with at least six different identifiable populations, with less heterogeneity of dendritic cells (DCs). Unsupervised modeling of developmental pathways combined with inference of gene regulatory networks suggested two major trajectories from common CCR2+ precursors resulting in CD11c+CD9+MMRintCD121b+ and CD11c-CD9intMMRhiCD121b- MP populations with unique transcription factors activities to coordinate downstream target molecules. The generation of both mature populations of colon MPs was impaired in germ-free mice, which coincided with the appearance of a dominant unique population of MPs, whereas the generation of DC populations in GF mice were unaffected or increased. Furthermore, the two major MP populations were clearly distinct from other populations regarding their gene expression profile, localization within the lamina propria, and ability to phagocytose macromolecules from the blood. These data uncover the diversity of intestinal myeloid cell populations, highlight the importance of microbiota on the unique developmental as well as anatomical and functional fates of colon MPs. We anticipate that these findings will form the basis for future studies on how tissue myeloid cell development is influenced either directly or indirectly by signals from endogenous microbiota in the intestine, as well as other tissues.

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Proteome of microparticles derived from Mycobacterium tuberculosis infected macrophages

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