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Abstract

Major histocompatibility complex (MHC) class II molecules are polymorphic cell surface
glycoproteins expressed only on the surface of the professional antigen presenting cells
and they play a crucial role in the adaptive immune responses. MHC class II molecules
undergo a very complex maturation process. They are synthesized in the endoplasmic
reticulum (ER), where
and  subunits of MHC class II molecules form heterodimers
and associate with the trimers of invariant chain (Ii), which targets the complex to late
endosomes.
Ii trimerization may be induced by the Ii transmembrane domain. Once the complex
arrives at the edosomes, Ii is proteolytically processed by any of the cysteine cathepsins
B, K, L, V and S undergoing a very complex regulation control. To enable biochemical
studies of the maturation and composition of the MHC class II molecules, its soluble
form, complexed with the two major forms, p31 and p41 of Ii, has been expressed in the
HEK293T cells at levels of 5 mg/l of the cell culture.
Here we analyzed the assembly and oligomeric state of the Ii and MHC class II
complexes lacking the transmembrane region. We have shown by in-vivo and in-vitro
analysis that Ii trimerizes also in the absence of the transmembrane part, prior to binding
of MHC class II dimers. The biochemical analysis supports the suggestion that the MHC
class II-Ii complexes are not necessarily trimers of trimer, as suggested before, but that
the Ii trimer can also be occupied by one or two MHC class II dimers. We also
demonstrate that p41 inhibitory fragment present on p41 form of Ii inhibited activity of
cathepsins L and V, but not cathepsins B and S.
Activity of cysteine cathepsins is regulated also by endogenous inhibitors stefins.
Therefore I was involved in biochemical and structural characterization of interactions
between cathepsins B and V and stefin A.