Upstream Regulators and Downstream Effectors of NADPH Oxidases as Novel Therapeutic Targets for Diabetic Kidney Disease

Oxidative stress has been linked to the pathogenesis of diabetic nephropathy, the complication of diabetes in the kidney. NADPH oxidases of the Nox family, and in particular the homologue Nox4, are a major source of reactive oxygen species in the diabetic kidney and are critical mediators of redox signaling in glomerular and tubulointerstitial cells exposed to the diabetic milieu. Here, we present an overview of the current knowledge related to the understanding of the role of Nox enzymes in the processes that control mesangial cell, podocyte and tubulointerstitial cell injury induced by hyperglycemia and other predominant factors enhanced in the diabetic milieu, including the renin-angiotensin system and transforming growth factor-β. The nature of the upstream modulators of Nox enzymes as well as the downstream targets of the Nox NADPH oxidases implicated in the propagation of the redox processes that alter renal biology in diabetes will be highlighted.     

On the name Stormbergia dangershoeki Butler

Imperfection in the formation of the name Stormbergia dangershoeki Butler is taken as an example so as to warn future taxon authors to repeat it.     

Cdc13 prevents telomere uncapping and Rad50-dependent homologous recombination.

Cdc13 performs an essential function in telomere end protection in budding yeast. Here, we analyze the consequences on telomere dynamics of cdc13-induced telomeric DNA damage in proliferating cells. Checkpoint-deficient cdc13-1 cells accumulated DNA damage and eventually senesced. However, these telomerase-proficient cells could survive by using homologous recombination but, contrary to telomerase-deficient cells, did so without prior telomere shortening. Strikingly, homologous recombination in cdc13-1 mec3, as well as in telomerase-deficient cdc13-1 cells, which were Rad52- and Rad50-dependent but Rad51-independent, exclusively amplified the TG(1-3) repeats. This argues that not only short telomeres are substrates for type II recombination. The Cdc13-1 mutant protein harbored a defect in its association with Stn1 and Ten1 but also an additional, unknown, defect that could not be cured by expressing a Cdc13-1- Ten1-Stn1 fusion. We propose that Cdc13 prevents telomere uncapping and inhibits recombination between telomeric sequences through a pathway distinct from and complementary to that used by telomerase.     

Glycosylation reduces the bioactivity of calcitonin

The biological significance of peptide hormone glycosylation is uncertain. To examine the effect of Asn-linked glycosylation on calcitonin's bioactivity we purified glycosylated calcitonin from a transplantable rat medullary thyroid carcinoma. Glycosylated calcitonin constituted 2.3% of the total extracted immunoreactive calcitonin. The structure of this peptide differed from nonglycosylated calcitonin only by the oligosaccharide modification of asparagine 3. Affinity of glycosylated calcitonin for lentil lectin indicated that the oligosaccharide was a complex processed form. In a standard in vivo bioassay glycosylated calcitonin had a markedly reduced hypocalcemic activity compared to nonglycosylated calcitonin, an effect most likely due to the presence of the oligosaccharide.     

A requirement for protein phosphorylation in regulating the meiotic and mitotic cell cycles in echinoderms

Populations of hormone-stimulated starfish oocytes and fertilized sea urchin eggs undergo synchronous meiotic and mitotic divisions. We have studied the requirement for protein phosphorylation during these events by testing the effects of 6-dimethylaminopurine (6-DMAP) upon the incorporation of [32P]orthophosphate. It was found that 6-DMAP blocked meiosis reinitiation and early cleavage and simultaneously inhibited protein phosphorylation, without changing the rate of [35S]methionine incorporation or pattern of protein synthesis. The protein, cyclin (54 kDa in starfish and 57 kDa in sea urchin), continues to be synthesized in the presence of 6-DMAP. This protein is destroyed at first and second cell cycles when 6-DMAP is added 30 min following fertilization but not when this drug is present before fertilization. Thus, cyclin breakdown does not depend on the completion of the nuclear events of M-phase, and its time of breakdown is set at an early step between fertilization and first cleavage. Using tubulin immunostaining, we found that 6-DMAP did not affect the cortical microtubules and resting female centrioles of prophase-arrested starfish oocytes, whereas it induced a precocious disappearance of spindle fibers when applied to hormone-stimulated oocytes. While an early addition of 6-DMAP precluded nuclear breakdown and spindle formation in both systems, a late treatment always allowed chromosome separation and centriole separation. Under these conditions pericentriolar tubulin persisted and could organize new spindles after the inhibitor was removed. It is suggested that (1) the assembly of cortical and centriolar-associated microtubules is not controlled by the same factors as spindle-associated tubulin; (2) specific proteins which are required for the cell to enter the following M-phase can become operative only via a process depending upon protein phosphorylation; (3) microtubule-associated kinases may play an important role in MPF function and spindle dynamics.     

Amino acid sequence of a mouse mucosal mast cell protease

The amino acid sequence has been determined of a mouse mucosal mast cell protease isolated from the small intestines of mice infected with Trichinella spiralis. The active protease contains 226 residues. Those corresponding to the catalytic triad of the active site of mammalian serine proteases (His-57, Asp-102, and Ser-195 in chymotrypsin) occur in identical positions. A computer search for homology indicates 74.3% and 74.1% sequence identity of the mouse mast cell protease compared to those of rat mast cell proteases I and II (RMCP I and II), respectively. The six half-cystine residues in the mouse mast cell protease are located in the same positions as in the rat mast cell proteases, cathepsin G, and the lymphocyte proteases, suggesting that they all have identical disulfide bond arrangements. At physiological pH, the mouse and rat mucosal mast cell proteases have net charges of +3 and +4, respectively, as compared to +18 for the protease (RMCP I) from rat connective tissue mast cells. This observation is consistent with the difference in solubility between the mucosal and connective tissue mast cell proteases when the enzymes are extracted from their granules under physiological conditions.     

A hypothesis on p34cdc2 sequestration based on the existence of Ca(2+)-coordinated changes in H+ and MPF activities during Xenopus egg activation [corrected]

The entry into, and exit from, mitosis are controlled by a universal M-phase promoting factor (MPF) composed of at least p34cdc2 and a cyclin. Embryonic systems are convenient for studying the association and dissociation of the active MPF complex because oocytes and eggs are naturally arrested at a specific point of the cell cycle until progression to the next point is triggered by a hormonal signal or sperm. In amphibians, eggs prior to fertilization are arrested at metaphase 2 of meiosis due to the presence of a stabilized MPF complex. Fertilization (egg activation) produces a transient increase in intracellular free Ca2+, a propagating Ca2+ wave, that specifically triggers the destruction of cyclin, leading to MPF inactivation and entry into the first embryonic inter-phase. We have recently shown that intracellular pH (pHi) variations in amphibian eggs, a large increase at fertilization and small oscillations during the embryonic cell cycle, were temporally and functionally related to the corresponding changes in MPF activity. In addition, the recent finding that the pHi increase at fertilization in Xenopus eggs is a propagating, Ca(2+)-dependent pH wave which closely follows the Ca2+ wave, together with the absence in the egg plasma membrane of pHi-regulating systems responsible for that pHi increase, suggest the existence of cortical or subcortical vesicles acidifying in the wake of the Ca2+ wave, thus producing the pH wave.(ABSTRACT TRUNCATED AT 250 WORDS)     

Demonstration that the leukocyte common antigen CD45 is a protein tyrosine phosphatase

It has been proposed on the basis of amino acid sequence homology that the leukocyte common antigen CD45 represents a family of catalytically active, receptor-linked protein tyrosine phosphatases [Charbonneau, H., Tonks, N. K., Walsh, K. A., & Fischer, E. H. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 7182-7186]. The present study confirms that CD45 possesses intrinsic protein tyrosine phosphatase (PTPase) activity. First, a mouse monoclonal antibody to CD45 (mAb 9.4) specifically eliminated, by precipitation, PTPase activity from a high Mr fraction containing CD45, prepared by gel filtration (Sephacryl S200) of a Triton X-100 extract of human spleen. Second, PTPase activity was demonstrated in a highly purified preparation of CD45 that was eluted with a high pH buffer from an affinity column, constructed from the same antibody. Third, on sucrose density gradient centrifugation, PTPase activity was only found in those fractions that contained CD45 as determined by Western analysis. When CD45 was caused to aggregate, first by reacting it with mAb 9.4 and then adding a secondary, cross-linking anti-mouse mAb, the PTPase activity shifted to the same higher Mr fractions that contained CD45. No shift in CD45 or PTPase was observed following addition of a control IgG2a. On this basis, it is concluded that CD45 is a protein tyrosine phosphatase.