The action radius of matrix metalloproteinases or MMPs is not restricted to massive extracellular matrix (ECM) degradation, it extends to the proteolysis of numerous secreted and membrane-bound proteins. Although many instances exist in which cells disintegrate, often in conjunction with induction of MMPs, the intracellular MMP substrate repertoire or degradome remains relatively unexplored. We started an unbiased exploration of the proteolytic modification of intracellularproteins by MMPs, using gelatinase B/MMP-9 as a model enzyme. To this end, multidimensional degradomics technology was developed by the integration of broadly available biotechniques. In this way, 100–200 MMP-9 candidate substrates were isolated, of which 69 were identified. Integration of these results with the known biological functions of the substrates revealed many novel MMP-9 substrates from the intracellular matrix (ICM), such as actin, tubulin, gelsolin, moesin, ezrin, Arp2/3 complex subunits, filamin B and stathmin. About 2/3 of the identified candidates were autoantigens described in multiple autoimmune conditions and in cancer (e.g. annexin I, nucleolin, citrate synthase, HMGB1, α-enolase, histidyl-tRNA synthetase, HSP27, HSC70, HSP90, snRNP D3). These findings led to the insight that MMPs and other proteases may have novel (immuno)regulatory properties by the clearance of toxic and immunogenic burdens of abundant ICM proteins released after extensive necrosis. In line with the extracellular processing of organ-specific autoantigens, proteolysis might also assist in the generation of immunodominant ‘neo-epitopes’ from systemic autoantigens. The study of proteolysis of ICM molecules, autoantigens, alarmins and other crucial intracellular molecules may result in the discovery of novel roles for proteolytic modification.

Multidimensional degradomics identifies systemic autoantigens and intracellular matrix proteins as novel gelatinase B/MMP-9 substrates.

The neuropathy associated with diabetes includes well documented impairment of axonal transport, a reduction in axon calibre and a reduced capacity for nerve regeneration. All of those aspects of nerve function rely on the integrity of the axonal cytoskeleton. Alterations in the axonal cytoskeleton in experimental diabetes include an insulin-dependent non-enzymatic glycation of actin that is reflected in increased glycation of platelet actin in the clinical situation. There is a reduced synthesis of mRNA for the isoforms of tubulin that are associated with nerve growth and regeneration and an elevated non-enzymatic glycation of peripheral nerve tubulin in both diabetic patients and diabetic animals. mRNAs for neurofilament proteins are selectively reduced in the diabetic rat and the post-translational modification of at least one of the neurofilament proteins is altered. There is some evidence that altered expression of isoforms of protein kinases may contribute to these changes.

The role of the axonal cytoskeleton in diabetic neuropathy

Although chronic inflammatory demyelinating polyneuropathy (CIDP) is presumed to be an autoimmune disorder, no neural antigen has been recognized as an immune target. We found that serum IgM from a patient with CIDP and an IgM paraprotein reacted with a 53-kd protein by Western blot analysis. Amino acid sequence analysis identified this protein as beta-tubulin. We then studied sera from 70 CIDP patients, 35 Guillain-Barre syndrome (GBS) patients, and 483 disease (amyotrophic lateral sclerosis, Alzheimer's disease, multiple sclerosis, diabetes, and other polyneuropathies) and normal controls for selective high-titer anti-beta-tubulin using ELISA methodology. Forty-two percent (30/70) of patients with CIDP had selective high titer IgM reactivity against beta-tubulin; 23% (16/70) had selective high-titer IgG reactivity against beta-tubulin. Overall, 57% of CIDP patients, 20% of GBS patients, and 2% of control patients had selective, high serum IgM or IgG anti-beta-tubulin reactivity. Selective high-titer serum anti-beta-tubulin antibodies occur in a majority of patients with CIDP but are rare in other chronic neuropathies or CNS disorders.

High‐titer selective serum anti‐β‐tubulin antibodies in chronic inflammatory demyelinating polyneuropathy

One of the cardinal manifestations of autoimmune disorders is the presence of autoantibodies and/or self reacting cells, however, the detection of these autoimmune activities is not necessarily associated with clinical findings. For example, several autoantibodies can be found in the sera of healthy family members of patients with autoimmune diseases1,2 and in sera of subjects with monoclonal gammopathies as well as in normal people 3,4

For Debate Parasitic Infection and Autoimmunity

Anti-histone autoantibodies react specifically with the B cell surface.

Sera from a total of 268 patients with protozoan, helminth, bacterial (leprosy and tuberculosis) infections or appropriate controls, were assayed for anti-tubulin antibodies in an indirect enzyme-linked immunosorbent assay (ELISA), using purified tubulin as antigen. Levels of serum anti-tubulin antibody were significantly elevated in 67% of patients with visceral leishmaniasis, in 60% of patients with cutaneous leishmaniasis, in 89% of patients with onchocerciasis, in 100% of patients with schistosomiasis, and in 94% of patients with leprosy. Little or no increase in anti-tubulin antibody levels was seen in sera from patients with malaria (Plasmodium vivax) or tuberculosis.

Howard Mk

Papers

Antibodies to tubulin in patients with parasitic infections.