Herrera E M, Ming M, Ortega-Barria E, Pereira M E
Department of Medicine, Tufts-New England Medical Center Hospitals, Boston, MA 02111.
Mol Biochem Parasitol. 1994 May;65(1):73-83. doi: 10.1016/0166-6851(94)90116-3.
Trypanosoma cruzi attaches and invades a large variety of mammalian cells by receptor-mediated interactions, one of them involving the binding of parasite trans-sialidase to host sialyl receptors. Three proteoglycan-deficient mutants of Chinese hamster ovary (CHO) cells were used to probe the role of host heparin and heparan sulfate glycosaminoglycans (GAG) in T. cruzi invasion. All three mutants supported adhesion and infection to a much lower extent than the parental CHO cells. One of the mutants, pgsD-677, did not express heparan sulfate while containing three- to four-fold excess chondroitin sulfate, yet the cell line was a poor substrate for T. cruzi adhesion. Proteoglycan-deficient cells obtained by inhibiting GAG synthesis in parental cells with p-nitrophenyl-beta-D-xyloside, were also poor hosts for T. cruzi invasion. Furthermore, digestion of parental cells with heparinase and heparitinase, two lyases that specifically depolymerize heparin and heparan sulfate, reduced the potential of the cells to support T. cruzi adhesion and growth. Lyases that digested chondroitin sulfate and other GAGs did not affect T. cruzi invasion. These results suggest that heparin/heparan sulfate epitopes are receptors for T. cruzi invasion. The corresponding counter-receptor on T. cruzi appears to be penetrin, a heparin-binding protein that promotes trypanosome penetration into cells. Purified penetrin caused agglutination of red blood cells, and the hemagglutination was exquisitely sensitive to heparin and heparan sulfate. However, sialic acid and sialyl compounds did not inhibit penetrin-induced hemagglutination. Recombinant penetrin competitively inhibited T. cruzi invasion of proteoglycan-containing parental cells, but not of proteoglycan-deficient mutants nor of heparitinase-treated cells. Furthermore, consistent with the sugar specificity of penetrin as a hemagglutinin, recombinant penetrin competed for trypanosome invasion of a CHO cell mutant (Lec2) that expresses heparan sulfate but not sialyl residues. Given that the release of sialic acid from the proteoglycan-deficient mutants further reduced T. cruzi invasion, as did the removal of heparan sulfate from the Lec2 mutant, and given that penetrin does not bind to sialic acid with high affinity, the results indicate that the penetrin-heparan sulfate pathway for T. cruzi invasion is distinct from the trans-sialidase-sialic acid route.
克氏锥虫通过受体介导的相互作用附着并侵入多种哺乳动物细胞，其中一种相互作用涉及寄生虫转唾液酸酶与宿主唾液酸受体的结合。利用中国仓鼠卵巢（CHO）细胞的三种蛋白聚糖缺陷型突变体来探究宿主肝素和硫酸乙酰肝素糖胺聚糖（GAG）在克氏锥虫入侵中的作用。与亲本CHO细胞相比，这三种突变体支持黏附和感染的程度要低得多。其中一个突变体pgsD - 677不表达硫酸乙酰肝素，而硫酸软骨素含量高出三到四倍，但该细胞系对克氏锥虫黏附而言是一种较差的底物。用对硝基苯基 - β - D - 木糖苷抑制亲本细胞中GAG合成获得的蛋白聚糖缺陷型细胞，对克氏锥虫入侵来说也是较差的宿主。此外，用肝素酶和硫酸乙酰肝素酶消化亲本细胞，这两种裂解酶能特异性地使肝素和硫酸乙酰肝素解聚，降低了细胞支持克氏锥虫黏附和生长的能力。消化硫酸软骨素和其他GAG的裂解酶不影响克氏锥虫的入侵。这些结果表明，肝素/硫酸乙酰肝素表位是克氏锥虫入侵的受体。克氏锥虫上相应的反受体似乎是穿膜蛋白，一种促进锥虫侵入细胞的肝素结合蛋白。纯化的穿膜蛋白可引起红细胞凝集，并且血凝反应对肝素和硫酸乙酰肝素极为敏感。然而，唾液酸和唾液酸化合物并不抑制穿膜蛋白诱导的血凝反应。重组穿膜蛋白竞争性抑制克氏锥虫对含蛋白聚糖的亲本细胞的入侵，但不抑制对蛋白聚糖缺陷型突变体或经硫酸乙酰肝素酶处理的细胞的入侵。此外，与穿膜蛋白作为血凝素的糖特异性一致，重组穿膜蛋白竞争克氏锥虫对表达硫酸乙酰肝素但不表达唾液酸残基的CHO细胞突变体（Lec2）的入侵。鉴于从蛋白聚糖缺陷型突变体中释放唾液酸进一步降低了克氏锥虫的入侵，从Lec2突变体中去除硫酸乙酰肝素也有同样效果，并且鉴于穿膜蛋白不以高亲和力结合唾液酸，结果表明克氏锥虫入侵的穿膜蛋白 - 硫酸乙酰肝素途径不同于转唾液酸酶 - 唾液酸途径。
