The E. coli SRP: preferences of a targeting factorDe Gier, Jan-Willem L; Valent, Quido A; Von Heijne, Gunnar; Luirink, Joen
doi: 10.1016/S0014-5793(97)00402-Xpmid: 9180256
Research on the targeting of proteins to the cytoplasmic membrane of E. coli has mainly focused on the so‐called ‘general secretory pathway’ (GSP) which involves the Sec‐proteins. Recently, evidence has been obtained for an alternative targeting pathway in E. coli which involves the signal recognition particle (SRP). The constituents of this SRP pathway in E. coli are homologous to those of the well‐characterized eukaryotic SRP pathway, which is the main targeting pathway for both proteins translocated across and inserted into the endoplasmic reticulum membrane. However, until recently, no clear function could be assigned to the SRP in E. coli. New studies point to an important role of the E. coli SRP in the assembly of inner membrane proteins.
Subunit interactions in the Escherichia coli protein translocase: SecE and SecG associate independently with SecYHomma, Takayuki; Yoshihisa, Tohru; Ito, Koreaki
doi: 10.1016/S0014-5793(97)00376-1pmid: 9180258
We used hexahistidine‐tagged SecE and SecY to study how the core subunits (SecY, SecE and SecG) of Escherichia coli protein translocase interact with each other. Detergent extracts were prepared from the plasma membranes and fractionated by Ni2+‐NTA agarose affinity binding. Although His6‐SecE, expressed in wild‐type cells, brought down both SecY and SecG, neither of them was brought down when the same protein was expressed in the secY24 mutant cells. His6‐SecY brought down both SecE and SecG, as expected. Interestingly, His6‐SecY24 was able to bring down SecG but not SecE. These results confirm our previous conclusion that the secY24 alteration impairs the SecY‐SecE interaction, and demonstrate that SecY and SecG can form a complex that does not contain SecE. Likewise, SecY‐SecE complex could be isolated from the secG‐deleted strain. The trimeric complex, in detergent extracts, dissociated at a critical temperature between 23 and 26°C, whereas the SecY‐SecE complex without SecG dissociated at a slightly lower temperature (20–23°C). We conclude that each of SecE and SecG independently binds to SecY, the central subunit of protein translocase, although the trimeric complex is more stable than the binary complexes.
Induction of glutathione synthetase by 1,10‐phenanthrolineSun, Yi
doi: 10.1016/S0014-5793(97)00380-3pmid: 9180259
The differential display (DD) was employed to identify the gene(s) responsible for 1,10‐phenanthroline (OP)‐induced apoptosis in murine tumor cells (Sun, Y., Bian, J., Wang, Y. and Jacobs, C. (1997) Oncogene 14, 385–393 [1]). An OP‐inducible gene was isolated which encodes mouse glutathione synthetase (GSS). The GSS mRNA level began to increase 6 h post OP treatment and remained at a high level thereafter up to 24 h tested. Induction of GSS was found not to be associated with p53 activation. No significant induction of DNA fragmentation was detected in two murine tumor lines upon GSS transfection. This is the first observation indicating that GSS is inducible rather specifically by a metal chelator and that induction of GSS, however, is not sufficient to induce apoptosis. It may merely reflect a cellular response to OP‐induced redox disturbance.
Deletion in the Z‐line region of the titin gene in a baby hamster kidney cell line, BHK‐21‐BiJäckel, Monika; Witt, Christian; Antonova, Olga; Curdt, Ingo; Labeit, Siegfried; Jockusch, Harald
doi: 10.1016/S0014-5793(97)00381-5pmid: 9180260
The gene for titin, a 4MDa myofibrillar protein, was analysed in golden hamster DNAs from different sources, using human cDNA probes and PCR. In the DNA from the BHK‐21‐Bi subline of baby hamster kidney cells, extended sequences coding for Z‐line associated domains were missing, indicating a deletion that renders titin non‐functional. These sequences were present in the original BHK‐21 line and in hamster DNAs. Our finding shows that, due to the absence of selective pressure on a gene's function, genomic deterioration can occur in a permanent cell line and can lead to a loss of overlapping DNA stretches in both autosomes.
Activation of Stat1 and subsequent transcription of inducible nitric oxide synthase gene in C6 glioma cells is independent of interferon‐γ‐induced MAP ...Nishiya, Tadashi; Uehara, Takashi; Edamatsu, Hiroki; Kaziro, Yoshito; Itoh, Hiroshi; Nomura, Yasuyuki
doi: 10.1016/S0014-5793(97)00383-9pmid: 9180263
Rat C6 glioma cells have been used to characterize molecular events involved in the regulation of inducible nitric oxide synthase (iNOS) gene expression stimulated by interferon‐γ (IFN‐γ) plus lipopolysaccharide (LPS). IFNs induce a signaling event which involves activation of Stat1 transcription factor. Previous studies have shown that IFNs also induce extracellular signal‐regulated kinase/mitogen‐activated protein kinase (ERK/MAPK) activation. However, the mechanisms by which IFNs stimulate MAPK activation remain elusive. Here we show that in C6 glioma cells, transiently expressing the dominant‐negative form of c‐Ha‐Ras (Asn‐17) abrogated IFN‐γ‐induced ERK1 and ERK2 activation. Furthermore, PD98059, a specific MEK1 inhibitor, also blocked this activation. These results indicate that p21
ras
and MEK1 are required for IFN‐γ‐induced ERK1 and ERK2 activation. Recent studies have reported that MAPK is responsible for serine phosphorylation of Stat1 which is required for Stat1's DNA binding and maximal transcriptional activity. Thus, we examined the role of the Ras‐MAPK pathway in Stat1 activation and subsequent iNOS induction in C6 glioma cells. Further experiments showed that neither Asn‐17 Ras expression nor concentrations of PD98059, which completely abrogated IFN‐γ‐induced ERK1 and ERK2 activation, affected Stat1 DNA binding activity or iNOS induction, indicating that the Ras‐MAPK pathway does not appear to be involved in the activation of Stat1 and subsequent iNOS induction in C6 glioma cells.
Uncoupling protein‐3: a new member of the mitochondrial carrier family with tissue‐specific expressionBoss, Olivier; Samec, Sonia; Paoloni-Giacobino, Ariane; Rossier, Colette; Dulloo, Abdul; Seydoux, Josiane; Muzzin, Patrick; Giacobino, Jean-Paul
doi: 10.1016/S0014-5793(97)00384-0pmid: 9180264
Brown adipose tissue (BAT) and skeletal muscle are important sites of nonshivering thermogenesis. The uncoupling protein‐1 (UCP1) is the main effector of nonshivering thermogenesis in BAT and the recently described ubiquitous UCP2 [1]has been implicated in energy balance. In an attempt to better understand the biochemical events underlying nonshivering thermogenesis in muscle, we screened a human skeletal muscle cDNA library and isolated three clones: UCP2, UCP3L and UCP3S. The novel UCP3 was 57% and 73% identical to human UCP1 and UCP2, respectively, highly skeletal muscle‐specific and its expression was unaffected by cold acclimation. This new member of the UCP family is a candidate protein for the modulation of the respiratory control in skeletal muscle.
Stimulation of human keratinocyte growth by alginate oligosaccharides, a possible co‐factor for epidermal growth factor in cell cultureKawada, Akira; Hiura, Nozomi; Shiraiwa, Masakazu; Tajima, Shingo; Hiruma, Masataro; Hara, Kenji; Ishibashi, Akira; Takahara, Hidenari
doi: 10.1016/S0014-5793(97)00386-4pmid: 9180265
Oligosaccharides, involved in regulation of plant developmental and defensive processes, were tested to determine their ability to enhance proliferation of human keratinocytes. A mixture of alginate oligosaccharides remarkably stimulated keratinocyte growth and [3H]thymidine uptake in the presence of epidermal growth factor (EGF). The activity was comparable to bovine pituitary extract, a common complement in keratinocyte culture, and additive on BPE‐induced stimulation. The most effective oligosaccharide in the mixture was identified and its chemical structure was determined. These findings demonstrate a novel activity of alginate oligosaccharide(s) in keratinocyte growth and suggest a possible co‐factor for EGF‐dependent stimulation in medium for keratinocytes.