To get a mechanistic insight in the regulation of Fas levels by lumican, we tested the possibility that Fas is regulated at the mRNA level. The RT-PCR results indicating equivalent mRNA levels eliminate the possibility that Fas mRNA is downregulated in the
Lum −/− mouse cornea. Consequently, the increase in Fas levels must be due to its regulation at the protein level. In the
Lum −/− mouse, FasL fails to elicit an increase in the receptor level, implying a lack of communication between the ligand and its receptor. We reasoned that lumican may be involved in concentrating and presenting FasL to its receptor Fas for appropriate induction, and we further investigated the binding of lumican to FasL.
11 Coimmunoprecipitation of lumican and FasL and solid state binding assay provide evidence for lumican-FasL interactions. Released from the membrane bound form of FasL (mFasL) by a putative metalloproteinase, the capacity of sFasL to induce apoptosis is much lower than that of mFasL, which is believed to be the functional form.
31 32 sFasL, in contrast, reportedly induces tumor cell
33 and hepatocyte apoptosis.
34 This discrepancy could arise from disparate effects of sFasL in tissues, where it may reside bound to different ECM components retaining much of the functions attributed to mFasL.
35 Thus, fibronectin was reported to bind soluble FasL (sFasL), which retained effector T cells at sites of inflammation and later induced T-cell apoptosis, healing, and return of tissue homeostasis.
35 36 Binding of FasL to fibronectin was proposed to result essentially in the oligomerization of FasL, further enhancing its biological activity.
36 In the eye, FasL is expressed by the corneal epithelium and endothelium.
37 The sFasL released in stroma may be similarly bound by the lumican present in abundance in the corneal stroma. It is likely, in the corneal stroma, that lumican is largely instrumental in retaining sFasL and increasing its biological efficacy.