Localization of atypical protein kinase C isoforms into lysosome-targeted endosomes through interaction with p62

An increasing number of independent studies indicate that the atypical protein kinase C (PKC) isoforms (aPKCs) are critically involved in the control of cell proliferation and survival. The aPKCs are targets of important lipid mediators such as ceramide and the products of the PI 3-kinase. In addition, the aPKCs have been shown to interact with Ras and with two novel proteins, LIP (lambda-interacting protein; a selective activator of lambda/iotaPKC) and the product of par-4 (a gene induced during apoptosis), which is an inhibitor of both lambda/iotaPKC and zetaPKC. LIP and Par-4 interact with the zinc finger domain of the aPKCs where the lipid mediators have been shown to bind. Here we report the identification of p62, a previously described phosphotyrosine-independent p56(lck) SH2-interacting protein, as a molecule that interacts potently with the V1 domain of lambda/iotaPKC and, albeit with lower affinity, with zetaPKC. We also show in this study that ectopically expressed p62 colocalizes perfectly with both lambda/iotaPKC and zetaPKC. Interestingly, the endogenous p62, like the ectopically expressed protein, displays a punctate vesicular pattern and clearly colocalizes with endogenous lambda/iotaPKC and endogenous zetaPKC. P62 colocalizes with Rab7 and partially with lamp-1 and limp-II as well as with the epidermal growth factor (EGF) receptor in activated cells, but not with Rab5 or the transferrin receptor. Of functional relevance, expression of dominant negative lambda/iotaPKC, but not of the wild-type enzyme, severely impairs the endocytic membrane transport of the EGF receptor with no effect on the transferrin receptor. These findings strongly suggest that the aPKCs are anchored by p62 in the lysosome-targeted endosomal compartment, which seems critical for the control of the growth factor receptor trafficking. This is particularly relevant in light of the role played by the aPKCs in mitogenic cell signaling events.     

Dimerization of recombinant horseradish peroxidase in a reversed micellar system

Recombinant horseradish peroxidase reactivated from E. coli inclusion bodies was studied in a reversed micellar system of AOT in octane. The ability of the recombinant enzyme, in contrast to native horseradish peroxidase, to form a dimeric structure was found. The existence of the dimer was proved by results of sedimentation analysis. Dimer/monomer ratio in the enzyme-containing micelles and dimer catalytic activity were found to depend on the substrate used (pyrogallol, guaiacol, o-dianisidine, o-phenylenediamine). Computer modelling was used to describe possible structures of the dimeric recombinant horseradish peroxidase.     

Properties of the catalytic center of a secretory 28kDa protein (1-cys peroxiredoxin) from rat olfactory epithelium

BACKGROUND: Percutaneous mitral valvuloplasty with the Inoue balloon is conventionally performed with double vascular access: arterial and venous. However, in patients with a good echogenic window it may be performed with venous access only and the procedure monitored by 2D-echocardiography and colour flow mapping. This should result in early ambulation and hospital discharge with reduced arterial complications. AIMS: To compare retrospectively the immediate results of percutaneous mitral valvuloplasty with the Inoue balloon in two groups of patients: Group I: venous access only (no arterial access, n = 102) and Group II: conventional double vascular access (arterial and venous access, n = 275). METHODS AND RESULTS: The baseline characteristics of the two groups were comparable for age, sex, clinical, echocardiographic, radiological and haemodynamic variables. The mitral valve area (Group I: 1.1 +/- 0.3 to 1.85 +/- 0.5 cm2 vs Group II: 1.05 +/- 0.2 to 1.85 +/- 0.5 cm2, P = ns) and transmitral gradient (Group I: 11 +/- 4 to 4.7 +/- 2 mmHg vs Group II: 12 +/- 4 to 4.8 +/- 2 mmHg, P = ns) before and after mitral valvuloplasty were not statistically different. A good immediate result, defined as mitral valve area > 1.5 cm2 and mean mitral gradient < 5 mmHg with mitral regurgitation < or = 2+ at the end of the procedure, was observed in 77% of the cases in the venous-only group and 79% in the double access group (P = ns). The incidence of severe mitral regurgitation (Grade III or IV) was not statistically significant. Procedural duration (71 +/- 24 min vs 109 +/- 26 min, P < 0.01), fluoroscopic time (12.5 +/- 5.5 min vs 18.5 +/- 6 min, P < 0.01) and hospital stay (2.8 +/- 1.5 days vs 4.8 +/- 2.6 days, P < 0.001) were significantly shorter in the venous-only group than in the conventional Inoue series. CONCLUSION: Single venous access balloon mitral valvuloplasty is as equally safe and effective as double vascular access. The additional advantages of single venous access are shorter procedural duration, fluoroscopic time and hospital stay. We recommend that it be performed by an experienced operator (minimum of 100 trans-septal punctures) in patients without major thoracic deformity and a good echogenic window.