http://hdl.handle.net/10033/116429 Wed, 22 May 2013 16:25:35 GMT 2013-05-22T16:25:35Z http://hdl.handle.net/10033/213569 Title: Internalization, phagolysosomal biogenesis and killing of mycobacteria in enucleated epithelial cells. Authors: de Souza Carvalho, Cristiane; Kasmapour, Bahram; Gronow, Achim; Rohde, Manfred; Rabinovitch, Michel; Gutierrez, Maximiliano Gabriel Abstract: Bacterial and parasitic intracellular pathogens or their secreted products have been shown to induce host cell transcriptional responses, which may benefit the host, favour the microorganism or be unrelated to the infection. In most instances, however, it is not known if the host cell nucleus is proximately required for the development of an intracellular infection. This information can be obtained by the infection of artificially enucleated host cells (cytoplasts). This model, although rather extensively used in studies of viral infection, has only been applied to few bacterial pathogens, which do not include Mycobacterium spp. Here, we investigate the internalization, phagosome biogenesis and survival of M. smegmatis in enucleated type II alveolar epithelial cells. Cytoplasts were infected with M. smegmatis, but the percentage of infection was significantly lower than that of nucleated cells. Scanning electron microscopy indicated that in both cells and cytoplasts, bacteria were internalized by a phagocytosis-like mechanism. Interestingly, phagosome fusion with lysosomes and mycobacterial killing were both more efficient in enucleated than in nucleated cells, a finding that may be correlated with the increased number of autophagic vesicles developed in cytoplasts. We provide evidence that although quantitative changes were observed, the full development of the infection, as well as mycobacterial killing did not require the presence of the host cell nucleus. Mon, 01 Aug 2011 00:00:00 GMT http://hdl.handle.net/10033/213569 2011-08-01T00:00:00Z http://hdl.handle.net/10033/116952 Title: Golgi-to-phagosome transport of acid sphingomyelinase and prosaposin is mediated by sortilin. Authors: Wähe, Anna; Kasmapour, Bahram; Schmaderer, Christoph; Liebl, David; Sandhoff, Konrad; Nykjaer, Anders; Griffiths, Gareth; Gutierrez, Maximiliano G Abstract: Sortilin, also known as neurotensin receptor 3 (NTR3), is a transmembrane protein with a dual function. It acts as a receptor for neuromediators and growth factors at the plasma membrane, but it has also been implicated in binding and transport of some lysosomal proteins. However, the role of sortilin during phagosome maturation has not been investigated before. Here, we show that in macrophages, sortilin is mainly localized in the Golgi and transported to latex-bead phagosomes (LBPs). Using live-cell imaging and electron microscopy, we found that sortilin is delivered to LBPs in a manner that depends on its cytoplasmic tail. We also show that sortilin participates in the direct delivery of acid sphingomyelinase (ASM) and prosaposin (PS) to the phagosome, bypassing fusion with lysosomal compartments. Further analysis confirmed that ASM and PS are targeted to the phagosome by sortilin in a Brefeldin-A-sensitive pathway. Analysis of primary macrophages isolated from Sort1(-/-) mice indicated that the delivery of ASM and PS, but not pro-cathepsin D, to LBPs was severely impaired. We propose a pathway mediated by sortilin by which selected lysosomal proteins are transported to the phagosome along a Golgi-dependent route during the maturation of phagosomes. Thu, 15 Jul 2010 00:00:00 GMT http://hdl.handle.net/10033/116952 2010-07-15T00:00:00Z