POBLACIÓN Y MUESTRA

D i s cu ss ed in detail in section 1.5.2, the cysteine pr ot e in a se s are very strongly imp licate d as being re spo ns ib le for the de gr ada ti on of invar ian t

chain from the immature class II a / p / I i trimeric com pl ex (De mo tz et al,

1994 ; Xu et al, 1994; Riese et al, 1996). Inv arian t chain degr ada ti on , as d eta il ed in section 1.6, is a p re req ui si te for class II pep ti de loading (Roche & Cre sswel l, 1991; Finesci et al, 1995). Cysteine pr ot e in a se r eq u ir em en t in such processes have been d em o ns tr at ed by inhib it or studies in which antigen pr ese nt at io n is im paired in B cells expo se d to such inhibitors, while overall proc ess ing function remains intact (Buus & Wer del in, 1987;

Dim ent , 1990, Reise et al, 1996). a / p / I i trime ric co m p le xe s have been

shown to accumulate in antigen p ro ces si ng co m pa rt m e nt s under such co nd i t i o n s (Blum & Cresswell, 1988; N ew co m b e & Cre sswell , 1993(b), W a r m e r d a m et al, 1996) with a co ns equ ent in hibition of class II pept ide l oa di ng and surface expression of class Il-ag com pl exe s (Neefjes & Ploegh,

1992) resu lting in the observed d o w nr egu lat io n in T cell stimu lation .

The aspartic proteinases have also been imp licate d as pl aying a role in li d e g r ad at io n (Nowell & Quaranta, 1985; Blum & Cres swe ll , 1988; Marie et al, 1994). From aspartic pr oteinase inhib it or studies, ca th eps in s D and E have been suggested as being re spo ns ib le for the initi atio n of li de gr ad at io n , while the cysteine p ro tein as e catalyse final release of li

f ra gm en ts from the a / p chains of class II (Marie et al, 1994). The use of

such inh ibito rs was shown to prevent forma ti on of class II a p ma turation.

The mo st likely cause of which however, is a lack of pep ti de su bstrate for class II binding rather than in hib ition of li degr adation. I n hib it io n of as p ar ti c prote ina se medi ated proc ess in g of end oc y to se d material and the r e s u l t i n g lack of peptides available for bin di ng would create an e n v i r o n m e n t in which class II will not mature. In addit ion, Reise et al

have p r ov id e d data showing that only cysteine prote ina se inh ib it or s prevent the fo rm at io n of mature MHC II co m p lex es (Reise et al, 1996). Fu rther evi de nc e against such a role for aspartic p ro tein as es exists in that li de gr ad at io n has been shown to remain un- im pa ir ed in the cat he ps in D kn o c k o u t mouse model (Saftig et al, 1995).

Ca th ep s in S has recently been strongly i m p li cat ed as a r e q u i re m e n t for li de gr ad ati on by studies ut ilising a novel, highly specific in h i b i t o r for this p r ot e in a se (Reise et al, 1996). In the pr ese nce of such an inh ib it or , a build

up of i m m at u re OC/p/Ii class II trimers has been shown to occur in a B cell line, with a co n co m it an t do w nr egu lat io n in p r es ent at io n fun ction. As such, the same effects as seen when using a cocktail of leupep tin and other cy steine p ro tei na se inhibitors are achie ved by using only the specific ca th ep si n S inhibitor. A 13Kd li d egr ada ti on pro du ct has been isolated

from in vi t ro cathepsin S a / p / I i digest studies which is not g en er at ed by other cys teine p ro teinases such as cathe ps ins B and H (Reise et al, 1996). Un like ca th eps in s B, D and E ( O ’Neil et al, u n p ub l i sh ed data), cat hep si n S

will not degrade the a and P chains of the MHC II trimer co m p le x and will

only attack the li component. When isolate d however, all three chain s are s u s ce pt ib le to cathepsin S mediated prote oly si s (Reise et al, 1996).

F ro m in vi t ro degradation studies, ca the ps in s D and B have bo th been

shown to degrade li from the class II a / p / I i trim er at a late en d o s o m a l pH of 4.5 ( O ’Neil et al, un pu bl ish ed data). In addition, the CC chain of the

trim er is also su sceptible to degra dat io n by the aspartic ca the ps in s, c at h ep s in E in particular. The deg radation prod ucts gener ated from the

a / p / I i tr i m er are distin ct for cathe ps in E, D and B ( O ’Neil et al,

u n p u b l i s h e d data) and appear to be disti nct from those ge ne r at ed by c at h ep s in S (Reise et al, 1996) suggesting perhaps, that the high degree of

sub str ate specificity of each cathe ps in pr o te in a se results in a unique d eg ra d at io n of li for each enzyme. In vi t ro studies designed to det er m ine the role of pr oteinases in the deg radation of li mu st take into acco unt the fact that such enzymes act in concert with other pr ote ina ses in thei r action. As such, studies concer ning a single enzym e in isolatio n may be limiting in elu cid at in g the precise role of ind iv id ual prote ina ses in li d egr ada ti on in vivo.