Mitochondrial rhodanese: membrane-bound and complexed activity

We have proposed that phosphorylated and dephosphorylated forms of the mitochondrial sulfurtransferase, rhodanese, function as converter enzymes that interact with membrane-bound iron-sulfur centers of the electron transport chain to modulate the rate of mitochondrial respiration (Ogata, K., Dai, X., and Volini, M. (1989) J. Biol. Chem. 204, 2718-2725). In the present studies, we have explored some structural aspects of the mitochondrial rhodanese system. By sequential extraction of lysed mitochondria with phosphate buffer and phosphate buffer containing 20 mM cholate, we have shown that 30% of the rhodanese activity of bovine liver is membrane-bound. Resolution of cholate extracts on Sephadex G-100 indicates that part of the bound rhodanese is complexed with other mitochondrial proteins. Tests with the complex show that it forms iron-sulfur centers when incubated with the rhodanese sulfur-donor substrate thiosulfate, iron ions, and a reducing agent. Experiments on the rhodanese activity of rat liver mitochondria give similar results. Taken together, the findings indicate that liver rhodanese is in part bound to the mitochondrial membrane as a component of a multiprotein complex that forms iron-sulfur centers. The findings are consistent with the role we propose for rhodanese in the modulation of mitochondrial respiratory activity.     

A growth-promoting factor existing on the body surface of rats

Effects of water-soluble matter adhering to rat hairs on fibroblasts were examined. The dialysate of the wash water of rat hairs significantly enhanced the cell proliferation of both diploid human dermal fibroblasts (DHDF) and diploid rat fibroblasts (DRDF). The cell growth-promoting activity was partially purified by a gel filtration column chromatography. The activity permeates through a ultrafiltration membrane (M.W. cut off: 500). Analyses of its chemical nature show that it is soluble in water, dimethyl sulfoxide or acetonitrile, insoluble in other organic solvents examined, stable to heat or pH shock, and resistant to a bacterial protease.     

Direct suppression of TCR-mediated activation of extracellular signal-regulated kinase by leukocyte protein tyrosine phosphatase, a tyrosine-specific phosphatase.

Leukocyte protein tyrosine phosphatase (LC-PTP)/hemopoietic PTP is a human cytoplasmic PTP that is predominantly expressed in the hemopoietic cells. Recently, it was reported that hemopoietic PTP inhibited TCR-mediated signal transduction. However, the precise mechanism of the inhibition was not identified. Here we report that extracellular signal-regulated kinase (ERK) is the direct target of LC-PTP. LC-PTP dephosphorylated ERK2 in vitro. Expression of wild-type LC-PTP in 293T cells suppressed the phosphorylation of ERK2 by a mutant MEK1, which was constitutively active regardless of upstream activation signals. No suppression of the phosphorylation was observed by LC-PTPCS, a catalytically inactive mutant. In Jurkat cells, LC-PTP suppressed the ERK and p38 mitogen-activated protein kinase cascades. LC-PTP and LC-PTPCS made complexes with ERK1, ERK2, and p38alpha, but not with the gain-of-function sevenmaker ERK2 mutant (D321N). A small deletion (aa 1-46) in the N-terminal portion of LC-PTP or Arg to Ala substitutions at aa 41 and 42 resulted in the loss of ERK binding activity. These LC-PTP mutants revealed little inhibition of the ERK cascade activated by TCR cross-linking. On the other hand, the wild-type LC-PTP did not suppress the phosphorylation of sevenmaker ERK2 mutant. Thus, the complex formation of LC-PTP with ERK is the essential mechanism for the suppression. Taken collectively, these results indicate that LC-PTP suppresses mitogen-activated protein kinase directly in vivo.     

Influence of External Osmotic Pressure on Water Permeability and Electrical Conductance of Chara Cell Membrane

Water permeability of the plasma membrane of a Characean internodalcell decreased with an increase in the osmotic pressure of theoutside of the cell, suggesting that the equivalent pore radiusof the water-filled pores becomes smaller with an increase inthe osmotic pressure. In contrast, the apparent membrane resistancedid not increase with an increase in the external osmotic pressure.These facts suggest that ions pass through the membrane mainlyvia pores other than those for bulk water flow. (Received October 22, 1986; Accepted May 22, 1987)     

Additive stimulation of hepatic putrescine production by glucagon and insulin after partial hepatectomy in rats

When rats received glucagon or insulin every 2 h after partial hepatectomy (Hx), hepatic putrescine content was increased above control levels at 6 and 12 h, respectively. When the two hormones were combined, the increased levels were additive. Hepatic ornithine decarboxylase activity was above control levels at 12 h after insulin treatment. Hepatic spermidine N1-acetyltransferase activity was enhanced at 6 h only when glucagon was dosed. Putrescine administration from 0 to 4 h or from 6 to 10 h increased hepatic DNA synthesis to similar levels 22 h after Hx. These results suggest that glucagon and insulin additively stimulate hepatic putrescine production after Hx. This may explain the cooperative stimulation of liver regeneration by both hormones.