Degradation of Human Laminin-332 by Candida Proteases

Objectives: Intact basement membrane is essential for maintaining the first line defence against pathogens. Candida proteases can degrade mouse laminin-332. We set out to study human laminin-332 degradation by 12 different Candida strains.

Methods: Human laminin-332 was produced by gingival keratinocyte cell-line (Tuija), labelled with 35S-methionine and isolated from the growth medium by immunoprecipitation. Candida strains used were: C. albicans (C. a.), C. dubliniensis (C. d), C. guilliermondii (C. gu), C. glabrata (C. gla), C. krusei (C. k) and C. tropicalis (C. t). Reference and clinical strains of each species were used. The cells were grown for 24 h, washed and disrupted by sonication. To obtain the cell free fraction the cells were removed from the growth media by filtration. Both the cell fractions and the 10 x concentrated cell free fractions were incubated with laminin-332 and proteolysis was assayed by fluorography. The effect of the incubation pH was tested by using two different pH values, pH 5.0 and pH 7.6.

Results: None of the Candida fractions degraded laminin-332 at pH 7.6. At pH 5.0 the cell fractions produced 100 kDa bands by both strains of C. d and C. t, clinical strains of C. a and C. gla, and reference strain of C. gu. The reference strain of C. k yielded 3 bands of 100-130 kDa size.The cell free fraction produced a band of 100 kDa size from both reference and clinical strains of C. a, C. d and C. t.

Conclusion: Degradation of human laminin-332 is pH dependent and differences exist between strains. The 100kDa sized particle could be the processed γ2-chain (γ2^') of laminin-332 (105kDa). Degradation can exert functional effects on the basement membrane, possibly aiding the invasion of Candida cells into the tissues. The results may thus cast some light on the virulence of Candida.