Secreted Dipeptidyl Peptidase IV Activity in the Dimorphic Fungal PathogenHistoplasma capsulatum

Abstract

Dipeptidyl peptidase type IV (DppIV) enzymes are broadly distributed phylogenetically and display diverse functions, including intercellular signaling, immunomodulation, protein maturation and processing, metabolism, and nutrient acquisition. We identified a secreted proteolytic activity in Histoplasma capsulatum effective toward DppIV-specific substrates. In order to determine the gene(s) that encodes this activity, we identified two putative DPPIV homologs (HcDPPIVA and HcDPPIVB) in H. capsulatum based on a homology search with Aspergillus fumigatus DppIV. Comparative sequence analysis revealed that HcDppIVA is similar to secreted DppIV enzymes, while HcDppIVB clusters with intracellular DapB-like enzymes. Unexpectedly, silencing of HcDPPIVA by RNA interference (RNAi) had no effect on secreted DppIV activity and an HcDPPIVA-null deletion mutant also showed no abrogation of secreted DppIV activity. In contrast, RNAi silencing of HcDPPIVB significantly reduced the level of secreted DppIV activity. RNAi silencing of HcDPPIVB in the HcDPPIVA-null mutant had no additional effect on secreted DppIV activity, indicating that HcDPPIVA does not contribute to secreted activity. RNAi silencing of HcDPPIVB did not affect the ability to kill a murine macrophage-like cell line, RAW 264.7, indicating that this gene is not required for infection of macrophages.

In vitro anti-diabetic activity of flavonoids and pheophytins from Allophylus cominia Sw . on PTP1B, DPPIV, alpha-glucosidase and alpha-amylase enzymes

“…Extracts of A. cominia at 30 µg/ml were incubated with DPPIV enzyme (purchased from Sigma Aldrich, UK, D7052) at 0.1 mg/ml for 30 min at 37 °C in a humidified atmosphere containing 5% CO 2 (Cooper and Woods, 2009). GlyoPro7-amido-4-methylcoumarin hydrobromide (10 µM, Sigma G2761, 25 mg) was then added and incubated for 30 min at 37 °C.…”

Fungal Mimicry of a Mammalian Aminopeptidase Disables Innate Immunity and Promotes Pathogenicity

“…CCL2 and CCR2 + cells promote resistance to this pathogen (Espinosa et al, 2014; Morrison et al, 2003) reminiscent of their roles with Blastomyces . In H. capsulatum , gene deletion of DppIVA did not cause a virulence defect (Cooper and Woods, 2009). However, DppIVB is responsible for most of the extracellular DppIV activity and has not been tested for a role in pathogenicity in vivo (Cooper et al, 2009).…”

Histoplasma mechanisms of pathogenesis - one portfolio doesn't fit all

“…Attempts to manipulate G186A and G217B in the lab have indicated differences in the efficiency of homologous recombination between the two strains. Whereas several gene deletion strains have been created through allelic replacement in the Panamanian background (G186A or G184A strains) (Woods et al, 1998;Sebghati et al, 2000;Tian & Shearer, 2002;Rappleye et al, 2004;Marion et al, 2006;Hwang et al, 2008;Hilty et al, 2011), only a limited number of gene knockout alleles exist in the NAm2 isolate G217B (Marion et al, 2006;Cooper & Woods, 2009). As a consequence, RNA interference (RNAi) has been adopted as a more practical means to deplete gene functions in Histoplasma (Rappleye et al, 2004) when efforts to delete genes through homologous recombination fail.…”