The Metalloprotease, Mpr1, Engages AnnexinA2 to Promote the Transcytosis of Fungal Cells across the Blood-Brain Barrier.
Eukaryotic pathogens display multiple mechanisms for breaching the blood-brain barrier (BBB) and invading the central nervous system (CNS). Of the fungal spp., that cause disease in mammals, only some cross brain microvascular endothelial cells which constitute the BBB, and invade the brain. Cryptococcus neoformans, the leading cause of fungal meningoencephalitis, crosses the BBB directly by transcytosis or by co-opting monocytes. We previously determined that Mpr1, a secreted fungal metalloprotease, facilitates association of fungal cells to brain microvascular endothelial cells and we confirmed that the sole expression of CnMPR1 endowed S. cerevisiae with an ability to cross the BBB. Here, the gain of function conferred onto S. cerevisiae by CnMPR1 (i.e., Sc<CnMPR1> strain) was used to identify targets of Mpr1 that might reside on the surface of the BBB. Following biotin-labeling of BBB surface proteins, Sc<CnMPR1>-associated proteins were identified by LC-MS/MS. Of the 62 proteins identified several were cytoskeleton-endocytosis-associated including AnnexinA2 (AnxA2). Using an in vitro model of the human BBB where AnxA2 activity was blocked, we found that the lack of AnxA2 activity prevented the movement of S. cerevisiae across the BBB (i.e., transcytosis of Sc<CnMPR1> strain) but unexpectedly, TEM analysis revealed that AnxA2 was not required for the association or the internalization of Sc<CnMPR1>. Additionally, the co-localization of AnxA2 and Sc<CnMPR1> suggest that successful crossing of the BBB is dependent on an AxnA2-Mpr1-mediated interaction. Collectively the data suggest that AnxA2 plays a central role in fungal transcytosis in human brain microvascular endothelial cells. The movement and exocytosis of Sc<CnMPR1> is dependent on membrane trafficking events that involve AnxA2 but these events appear to be independent from the actions of AnxA2 at the host cell surface. We propose that Mpr1 activity promotes cytoskeleton remodeling in brain microvascular endothelial cells and thereby engages AnxA2 in order to facilitate fungal transcytosis of the BBB.
['Na Pombejra|Sarisa|S|', 'Salemi|Michelle|M|', 'Phinney|Brett S|BS|', 'Gelli|Angie|A|']
D000123:Acetyltransferases D017306:Annexin A2 D001692:Biological Transport D001812:Blood-Brain Barrier D001921:Brain D002460:Cell Line D002465:Cell Movement D002478:Cells, Cultured D002490:Central Nervous System D003455:Cryptococcus neoformans D003599:Cytoskeleton D003600:Cytosol D004705:Endocytosis D042783:Endothelial Cells D015966:Gene Expression Regulation, Fungal D006790:Host-Parasite Interactions D006801:Humans D008590:Meningoencephalitis D045726:Metalloproteases D012441:Saccharomyces cerevisiae D029701:Saccharomyces cerevisiae Proteins D053719:Tandem Mass Spectrometry D057900:Transcytosis
metalloprotease Mpr1 AnnexinA2 fungal cells blood-brain barrier Cryptococcus neoformans Saccharomyces cerevisiae mass spectrometry