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A Katz

Publications and source records attributed to A Katz.

At least 181 records · Page 10Linked to original sources
PubMed1995

Insulin-mediated activation of glycogen synthase in isolated skeletal muscle: role of mitochondrial respiration.

Activation of glycogen synthase (GS) by insulin in isolated, aerated muscle was abolished when oxygen was replaced by nitrogen. Reoxygenation of the muscle after prior exposure to nitrogen restored the insulin effect. In the presence of cyanide (inhibitor of mitochondrial respiration) insulin did not activate GS. These data demonstrate that adequate mitochondrial respiration is required for insulin-mediated activation of GS.

Adenosine Triphosphate↗
PubMed1995

On-site catheterization laboratory and prognosis after acute myocardial infarction. Israeli Thrombolytic Survey Group.

BACKGROUND: Since the introduction of thrombolytic therapy for patients with acute myocardial infarction, the use of coronary angiography has substantially increased. We sought to determine whether the presence of on-site coronary angiographic facilities influenced the utilization of coronary procedures in patients with acute myocardial infarction hospitalized in Israel's coronary care units. METHODS: A prospective survey was conducted in January and February 1992 in the 25 coronary care units operating in Israel, 15 of which had on-site catheterization facilities. Data on demographics, clinical features, thrombolytic therapy, and the type of coronary diagnostic or therapeutic procedures performed during the current in-hospital stay were recorded. Mortality, both in-hospital and 1 year after discharge, was assessed for all patients in the survey. RESULTS: One thousand fourteen consecutive patients with acute myocardial infarction were hospitalized during the survey, 307 (30%) of whom were admitted to 10 coronary care units without and 707 of whom were treated in hospitals with on-site coronary angiography facilities. Demographic and baseline characteristics were similar in both groups. Thrombolytic therapy was provided equally (46%) to patients admitted to hospital with and without catheterization laboratories. Patients admitted to hospitals with these laboratories underwent coronary angiography (26%) and percutaneous transluminal angioplasty and/or coronary artery bypass grafting (12%) in greater numbers than counterparts admitted to hospitals without such laboratories (10% and 5%, respectively). Hospital and cumulative 1-year mortality rates were 11% and 18%, respectively, in patients admitted to hospitals with on-site catheterization facilities vs 10% and 17%, respectively, in the patient group admitted to the other hospitals. Patients receiving thrombolytic therapy had similar hospital mortality rates unrelated to the availability of coronary catheterization laboratories. CONCLUSION: This national survey showed that the availability of invasive coronary facilities led to increased use of diagnostic and therapeutic coronary procedures among patients with acute myocardial infarction. There was no difference in hospital or 1-year mortality rates in patients admitted to hospitals with or without on-site coronary angiographic facilities.

Aged↗
PubMed1995

A segment of the C-terminal half of the G-protein beta 1 subunit specifies its interaction with the gamma 1 subunit.

The beta and gamma subunits of the heterotrimeric guanine nucleotide binding (G protein) act as a dimer and directly regulate various signal transduction pathways. By using cotransfection assays, we tested the ability of several beta gamma combinations to activate inositol phospholipid-specific phospholipase C (PI-PLC)-beta 2. Our findings indicate that only beta gamma combinations that form dimers will activate PI-PLC-beta 2. Since G beta 1 interacts with G gamma 1, while G beta 2 cannot, chimeras between G beta 1 and G beta 2 were used to identify the regions in beta 1 that determine its specific association with gamma 1. Our evidence demonstrates that a chimera between beta 2 and beta 1 that contains the C-terminal 173 amino acids of beta 1 can interact and activate PI-PLC-beta 2 with gamma 1. Chimeras that contain portions of the beta 1 C-terminal region display a weaker association with gamma 1. Furthermore, the contribution of each of these regions depends on the sequence context of each chimeric protein. However, the segment between residues 210 and 293 of beta 1 consistently plays a critical role in specifying association with gamma 1.

Amino Acid Sequence↗
PubMed1995

Functional analysis of a dominant negative mutant of G alpha i2.

The key event in receptor-catalyzed activation of heterotrimer G proteins is binding of GTP, which leads to subunit dissociation generating GTP-bound alpha subunits and free beta gamma complexes. We have previously identified a mutation that abolished GTP binding in G alpha o (S47C) and demonstrated that the mutant retained the ability to bind beta gamma and could act in a dominant negative fashion when expressed in Xenopus oocytes (Slepak, V.Z., Quick, M.W., Aragay, A.M., Davidson, N., Lester, H.A., and Simon, M.I. (1993) J. Biol. Chem. 268, 21889-21894). In the current work, we investigated the effects of the homologous mutant of G alpha i2 (S48C) upon signaling pathways reconstituted in transiently transfected COS-7 cells. We found that expression of the G alpha i2 S48C mutant prevented stimulation of phospholipase C (PLC) beta 2 by free beta gamma subunit complexes. This effect of G alpha i S48C was not readily reversible in contrast to the inhibitory effect of wild-type G alpha i2, which could be reversed upon activation of the cotransfected muscarinic M2 receptor, presumably by release of beta gamma from the G protein heterotrimer. Coexpression of G alpha i S48C or the wild-type G alpha i2 also dramatically decreased G16-mediated stimulation of PLC by C5a in the cells transfected with cDNAs encoding C5a receptor and G alpha 16. Activation of PLC via endogenous Gq or G11 in the presence of alpha 1C adrenergic receptors was similarly attenuated by coexpression of G alpha i or G alpha i S48C. Pertussis toxin treatment of the transfected cells enhanced the inhibition of the receptor-stimulated PLC by wild-type G alpha i subunits but did not influence the effects of the dominant negative mutant. The enhancement of the wild-type G alpha i inhibitory effect by pertussis toxin can be explained by stabilization of G alpha i binding to beta gamma as a result of ADP-ribosylation, while G alpha i S48C mutant binds beta gamma irreversibly even without pertussis toxin treatment. Therefore, a feasible mechanism to rationalize the attenuation of the G alpha 16 and Gq/11-mediated activation of PLC by cotransfected G alpha i is the competition between G alpha i and G alpha 16 or Gq/11 for the beta gamma complexes, which are necessary for the G protein coupling with receptors. These experiments provide new evidence for the role of beta gamma in the integration of signals controlling phosphoinositide release through different G alpha families.

Adenosine Diphosphate Ribose↗
PubMed1995

Rapid activation of glycogen synthase and protein phosphatase in human skeletal muscle after isometric contraction requires an intact circulation.

The effects of isometric contraction (66% of maximal force) and recovery on glycogen synthase fractional activity (GSF) in human skeletal muscle have been studied. Biopsies were taken from the quadriceps femoris muscle at rest, at fatigue and 5 min postexercise on two occasions: after one of the contractions, the circulation to the thigh was occluded during the 5 min recovery (OCC), and after the other contraction, the circulation was intact (control, CON). During CON, GSF decreased from (mean +/- SE) 0.34 +/- 0.05 at rest to 0.24 +/- 0.02 at fatigue and then increased to 0.74 +/- 0.04 at 5 min postexercise; corresponding values for OCC were 0.37 +/- 0.04, 0.25 +/- 0.04 and 0.48 +/- 0.05 (P < 0.001 vs. CON for 5 min postexercise only). Compared with the value at fatigue, protein phosphatase activity (PP) increased by 79 +/- 16% during CON recovery (P < 0.01), whereas no change was observed during OCC recovery. Uridine diphosphate glucose increased by approximately 2.5-fold at fatigue, remained elevated during OCC recovery, but reverted to the preexercise level during CON recovery (P < 0.001 vs. OCC recovery). Glucose 6-P increased approximately 5-fold at fatigue and was higher at 5 min postexercise in OCC vs. CON recovery (8.6 +/- 1.5 vs. 4.1 +/- 0.9 mmol/kg dry wt; P < 0.01). It is concluded that the rapid increase in GSF after intense exercise with an intact circulation may be at least partly attributed to an increase in the specific activity of PP. The increase in GSF during recovery in OCC may be at least partly attributed to the high glucose 6-P content in vivo, which enhances the substrate suitability of GS for PP. Thus, separate mechanisms exist for the activation of PP and GS during recovery from intense short term exercise.

Adult↗
PubMed1995

A prospective randomized evaluation of three schedules of mesna administration in patients receiving an ifosfamide-containing chemotherapy regimen: sustained efficiency and simplified administration.

Chemotherapy with oxazaphosphorines, such as ifosfamide, is often limited by unacceptable urotoxicity. Without uroprotection hemorrhagic cystitis becomes dose-limiting. Mesna, a thiol compound, is a drug able to bind the toxic metabolites, forming nontoxic compounds in the urine. A total of 122 patients were enrolled in this study and 228 chemotherapy cycles with an ifosfamide-containing regimen were performed (225 evaluable). Mesna was given at the same total dose as the ifosfamide in all arms. On arm A, mesna was given i. v. in equal doses 15 min before and 4 h and 8 h following the ifosfamide dose. On arm B, mesna was given in three equivalent doses 15 min before (i.v.) and 4 h (i.v.) and 8 h (p.o., double dose) following ifosfamide. On arm C, mesna was given i.v. in two equal doses given 15 min before and 4 h following. The incidence of urotoxicity was very low (lower than 15%) in the three arms, 0% in A, 1.36% in B and 2.70% in C. All three arms were equally efficient. Schedule C was considered superior to the others, since it was equally effective, simpler and more convenient.

Adolescent↗
PubMed1995

An interlocking auricular composite graft.

BACKGROUND: Full-thickness defects of the nasal alar rim are relatively common following Mohs micrographic surgery for the treatment of long-standing or recurrent skin tumors. Composite grafts provide an excellent cosmetic and functional alternative for the repair of such defects. OBJECTIVE: A useful technique of auricular composite graft placement for reconstruction of full-thickness nasal alar rim defects is described. METHODS: The cartilaginous portion of the graft is extended beyond the borders of the soft tissue defect so that two cartilaginous pegs frame the lateral aspects of the graft. These pegs are then inserted into pockets prepared within the alar tissue of both sides of the defect, such that the graft interlocks with its recipient bed. A series of diagrams as well as a set of photographs from a representative case are provided, along with accompanying commentary, so as to enable the surgeon to incorporate this technique easily into his/her practice. CONCLUSION: The interlocking auricular composite graft technique permits increased graft stability, with decreased shearing forces of the graft over its recipient bed, and a larger surface area for revascularization, resulting in an increased probability of graft survival. This technique provides an elegant single stage alternative to current reconstructive techniques for full-thickness nasal alar rim defects measuring less than 1.5 cm in diameter.

Bandages↗
PubMed1995

Evaluation of a new rate adaptive ventricular pacemaker controlled by double sensors.

UNLABELLED: The LEGEND-PLUS, a new rate adaptive pacemaker that combines activity and minute ventilation sensing for automatic rate adaptation was implanted in the right ventricle (VVIR) in 11 patients (mean age 62 +/- 9 years). Initial programming was performed using the Programmer Exercise Protocol (a 3-minute walk). This programming was evaluated by treadmill tests, up-stairs and down-stairs walking, and Holter recordings. RESULTS: Following the final programming of LEGEND-PLUS, the mean upper activity rate was 102 +/- 7 beats/min (range 90-120 beats/min), while the mean upper minute ventilation rate was 125 +/- 16 beats/min (range 100-150 beats/min). The mean rate responses during the exercise protocol and the final programming in minute ventilation and activity sensing modes were 5.4 +/- 2.3 (range 1-9), versus 4 +/- 2.4 (range 1-8; P < 0.01) and 7.6 +/- 1.1 (range 5-9), versus 7.5 +/- 0.8 (range 6-9; P = 0.8), respectively. In the combined sensing mode, the acceleration rate was identical to the activity rate response and the deceleration rate mimicked the minute ventilation. CONCLUSION: Dual sensor VVIR pacemakers have the potential to improve rate adaptation to exercise. The rate response to exercise in patients fitted with activity and minute ventilation sensors, VVIR pacemakers closely mimics the physiological rate response.

Adult↗
PubMed1995

Inhibition of superoxide production in human neutrophils by combinations of heparin and thrombolytic agents.

OBJECTIVE: To investigate the effect of heparin and thrombolytic agents on superoxide generation by human neutrophils, as inhibition of superoxide production may have a role in reducing ischaemia and reperfusion injury. METHODS: Neutrophil superoxide production stimulated by phorbol myristate acetate (PMA), opsonised zymosan, or formyl methionyl leucyl phenylalanine (FMLP) was measured as the superoxide dismutase inhibitable reduction of acetyl ferricytochrome c by a microtitre plate technique. RESULTS: Heparin, at concentrations of 0.5-500 U/ml, caused a gradual inhibition of superoxide production stimulated by PMA, opsonised zymosan, or FMLP. Tissue plasminogen activator was more potent than heparin in inhibiting superoxide production induced by opsonised zymosan or FMLP, but it did not affect the activity stimulated by PMA. Streptokinase or urokinase had no effect on superoxide production. When heparin was used in combination with tissue plasminogen activator, streptokinase, or urokinase at their therapeutic concentrations there was a significant inhibition of superoxide generation (70%, 30%, and 25%, respectively). The therapeutic concentrations of tissue plasminogen activator alone caused a reduction of 40% of neutrophil superoxide production. When tissue plasminogen activator and streptokinase were both added to neutrophils, however, a synergistic inhibition of 80% was achieved. CONCLUSIONS: The inhibition of super oxide generation by these drug combinations may explain the limited inflammatory response and reduction of reperfusion injury observed in patients receiving thrombolytic treatment.

Depression, Chemical↗
PubMed1995

Splanchnic and muscle metabolism during exercise in NIDDM patients.

To characterize splanchnic and muscle metabolism during exercise in non-insulin-dependent diabetes mellitus (NIDDM), eight male nonobese patients and seven healthy control subjects (CON) were studied during 40 min of bicycle exercise at 60% of maximal oxygen uptake. Biopsies were obtained from the quadriceps femoris muscle at rest and immediately after exercise. Arterial glucose concentration in NIDDM had declined by 10% (P < 0.01) at the end of exercise, whereas in CON it had risen by 21% (P < 0.05). Leg glucose uptake rose from 0.19 +/- 0.06 mmol/min at rest to 2.25 +/- 0.61 mmol/min at the end of exercise in NIDDM and from 0.13 +/- 0.05 to 1.17 +/- 0.34 mmol/min in CON. Splanchnic glucose output increased from 0.52 +/- 0.06 to 2.37 +/- 0.26 mmol/min in NIDDM and from 0.79 +/- 0.12 to 2.44 +/- 0.38 mmol/min in CON. Leg lactate output during exercise was twofold higher in NIDDM. Muscle contents of lactate and glycogen were similar in both groups at rest, whereas after exercise lactate tended to be higher (19.5 +/- 1.7 vs. 12.7 +/- 5.9 mmol/kg dry wt) and glycogen lower (154 +/- 35 vs. 251 +/- 41 mmol glucosyl units/kg dry wt) in NIDDM. Whole body respiratory exchange ratio during exercise was higher in NIDDM (0.84 +/- 0.02 vs. 0.78 +/- 0.02, P < 0.05). Exercise-induced changes in other muscle metabolites were similar in NIDDM and CON. These data indicate that the decline in blood glucose during exercise in nonobese NIDDM is due to enhanced peripheral glucose utilization rather than to an attenuated increase in splanchnic glucose output.

Adult↗
PubMed1995

Multiple regions of G alpha 16 contribute to the specificity of activation by the C5a receptor.

The C5a chemoattractant factor receptor, when expressed in COS-7 cells, can stimulate phosphoinositide-specific phospholipase C activity through the activation of the G16 isoform of the heterotrimeric G protein, but not through the G11 isoform. To identify the regions of the G alpha 16 subunit protein that are responsible for its activation by the C5a receptor, a series of chimeras between G alpha 16 and G alpha 11 were constructed and tested for their ability to be activated by the C5a receptor. Co-transfection experiments with chimeras in which the carboxyl-terminal regions of G alpha 11 were replaced with the corresponding regions of G alpha 16 demonstrated that changes in the carboxyl terminus, e.g., replacement of 134 amino acids, were not sufficient to confer receptor specificity. An additional segment encompassing residues 220-240 of G alpha 16 was required to confer C5a-induced activation. Testing of a reciprocal series of chimeras composed of G alpha 16 sequences at the amino terminus and G alpha 11 sequences at the carboxyl terminus revealed that certain sequences extending from the amino terminus to amino acid 209 of G alpha 16 were sufficient to endow the chimera with much of the specificity for C5a-induced activation. These results suggest that receptor specificity may involve specific conformations of the G protein stabilized by concerted interactions of multiple amino acid sequences distributed throughout the G alpha protein.

Animals↗
PubMed1995

Oxidized glutathione as a marker of ischemia reperfusion associated with single lung transplantation.

BACKGROUND: Previous data have shown that glutathione (GSH), an endogenous antioxidant, is converted to its oxidized form (GSSG) after oxidative ischemia-reperfusion events. As GSSG is toxic to cells and is extruded through an active mechanism dependent on intracellular GSSG levels, substance appears in plasma. STUDY DESIGN: Single lung transplantation was performed upon 18 puppies, 3 to 5 kg, with a two hour ischemic time for the donor lung before reimplantation. Recipient animal plasma was obtained after anesthesia induction, pulmonary artery ligation, recipient pneumonectomy, reestablishment of blood flow to the donor lung, completion of transplant, and one, two, and three hours postoperatively. Donor lung bronchoalveolar lavage fluid (BALF) was obtained at the time of harvest, after perfusion of the donor lung with EuroCollins, immediately pretransplant, and after completion of the vascular anastomoses. Oxidized and total GSH levels in plasma and BALF samples were determined by a spectrophotometric assay. RESULTS: After reimplantation of the ischemic donor lung, there was a statistically significant increase in both GSSG and GSH in plasma samples, and a statistically significant increase in GSSG in the BALF. CONCLUSIONS: Compartmental differences between arterial and venous plasma, as well as the increase in GSSG in the BALF, implicated the lung that was transplanted as the source of oxygen free radical generation and GSSG release. Plasma GSSG levels seem to provide a sensitive, noninvasive, repeatable measure of ongoing tissue response and oxygen free radical production.

Animals↗
PubMed1995

Induction of release of secretory nonpancreatic phospholipase A2 from human articular chondrocytes.

OBJECTIVE: Secretory nonpancreatic phospholipase A2 (sPLA2) is a known inducer/promoter of the inflammatory process in the joints. It correlates with disease activity in adult and juvenile rheumatoid arthritis. Synovial fluids contain high concentrations of sPLA2. We discovered that human articular cartilage contains large quantities of sPLA2 and that culture chondrocytes constitutively synthesize and release sPLA2. To test the mechanism controlling the release of sPLA2, we exposed cultured human articular chondrocytes to cytokines and other agents, known to induce sPLA2 in other cells. METHODS: Chondrocytes obtained from cartilage of normal appearance from rheumatoid and osteoarthritic joints, and from normal, neonatal joints were compared to rabbit articular chondrocytes. Radiolabeled Escherichia coli derived phospholipid assay and ELISA technique using monoclonal antibodies against recombinant human synovial type sPLA2 were employed. The cells were grown as monolayers as well as in alginate beads. RESULTS: Human articular chondrocytes from both arthritic and neonatal joints released sPLA2 constitutively but could not be further stimulated with interleukin-1 beta (IL-1 beta), tumor necrosis factor alpha (TNF-alpha), IL-6, oncostatin M, lipopolysaccharide (LPS), or forskolin. Marked stimulation was observed when the cells were exposed to 8-bromo cyclic adenosine monophosphate (cAMP). Growing the cells as monolayers or in alginate beads did not change the results. In contrast to human cells, rabbit chondrocytes responded to IL-1 beta and IL-1/TNF, but not to TNF-alpha alone, with a very marked increase in extracellular sPLA2 activity. CONCLUSION: Human articular chondrocytes synthesize and constitutively release sPLA2. Such continuous release is most probably responsible for the high concentration of sPLA2 in articular cartilage and may be the source of synovial fluid sPLA2. To our knowledge, human articular chondrocytes are the only sPLA2 producing cells tested to date that do not respond to cytokine stimulation with increased sPLA2 activity; yet enhancement was seen with 8-bromo cAMP. It seems therefore that, human articular chondrocytes possess signalling mechanisms for the release of sPLA2 unlike those from other mammalian cells. The significance of this observation remains to be elucidated.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1994

Hydrocortisone differentially affects the ability of murine stromal cells and human marrow-derived adherent cells to promote the differentiation of CD34++/CD38- long-term culture-initiating cells.

Very primitive human hematopoietic progenitor cells are identified indirectly by their ability to give rise to clonogenic progenitors in the presence of either human or murine stromal cells. These long-term culture-initiating cell (LTC-IC) assays are usually performed in the presence of hydrocortisone based on the initial observation that hydrocortisone was required for prolonged hematopoiesis in standard long-term bone marrow cultures. In this report, we investigated the role of hydrocortisone in LTC-IC assays initiated with CD34++/CD38- cells seeded onto either human bone marrow LTC-derived adherent cells or a murine marrow-derived stromal cell line, MS-5. It was found that weekly addition of hydrocortisone to the cultures reduced the frequency of LTC-IC (from 1/5 to 1/20) calculated from limiting dilution experiments and also reduced fivefold to 10-fold the number of their progeny clonogenic cells detected after 4 to 5 weeks. In contrast, the frequency and differentiative potential of CD34++/CD38- grown in the presence of human marrow feeders was unaltered by the addition of glucocorticoids. Data are consistent with the hypothesis that hydrocortisone inhibited LTC-IC differentiation by downregulating the expression of a synergistic factor produced by MS-5 cells. (1) In the absence of hydrocortisone, the number of clonogenic progenitors generated by LTC-IC was much higher in cultures seeded on MS-5 than in cultures seeded on human marrow adherent cells, which was also true when cytokines were added to the cocultures. However, based on the phenotype of the colonies, progenitors produced in MS-5 cocultures were more mature than those generated on human marrow adherent cells. (2) Hydrocortisone counteracted the stimulatory effect of recombinant human cytokines (interleukin-3, interleukin-6, and steel factor) in assays performed on MS-5 but not on human marrow feeders. (3) Hydrocortisone led to a 50% decrease in the numbers of colony-forming units-granulocyte-macrophage found in methycellulose colony assays of CD34++/CD38- cells performed in the presence of MS-5 cells. Taken together, our results indicate that hydrocortisone acts differently on a murine stromal cell line and on marrow-derived human stromal cells and may suppress the expression by MS-5 cells of an activity selectively promoting amplification of clonogenic cells derived from primitive LTC-IC.

ADP-ribosyl Cyclase↗
PubMed1994

[Do on-site coronary angiographic facilities influence management and prognosis of acute myocardial infarction?].

Whether the presence of on-site coronary angiographic facilities (CAF) influences the use of invasive coronary procedures and the outcome of acute myocardial infarction in coronary care units was studied. A prospective survey was conducted early in 1992 when 1014 consecutive patients with acute infarction were admitted. Of them 707 (70%) were admitted to coronary care units of hospitals with, and 307 (30%) without CAF. Thrombolytic therapy was given to 46% in both groups. Those admitted to hospitals with CAF underwent more coronary angiographies (26%) and percutaneous transluminal angiography and/or coronary artery bypass grafting (12%) than those without CAF (10 and 5%, respectively--p < 0.005). In-hospital and 1-year mortality were 11 and 18% respectively in those with CAF, vs 10 and 11%, respectively, in those without CAF. All patients treated with a thrombolytic agent had similar mortality, unrelated to the availability of CAF. The survey showed that the availability of on-site coronary angiography facilities led to greater use of invasive coronary procedures in cases of acute myocardial infarction, but there was no significant difference in mortality.

Angioplasty, Balloon, Coronary↗
PubMed1994

Binding of beta gamma subunits of heterotrimeric G proteins to the PH domain of Bruton tyrosine kinase.

Bruton tyrosine kinase (Btk) has been implicated as the defective gene in both human and murine B-cell deficiencies. The identification of molecules that interact with Btk may shed light on critical processes in lymphocyte development. The N-terminal unique region of Btk contains a pleckstrin homology domain. This domain is found in a broad array of signaling molecules and implicated to function in protein-protein interactions. By using an in vitro binding assay and an in vivo competition assay, the pleckstrin homology domain of Btk was shown to interact with the beta gamma dimer of heterotrimeric guanine nucleotide-binding proteins (G proteins). A highly conserved tryptophan residue in subdomain 6 of the pleckstrin homology domain was shown to play a critical role in the binding. The interaction of Btk with beta gamma suggests the existence of a unique connection between cytoplasmic tyrosine kinases and G proteins in cellular signal transduction.

Agammaglobulinaemia Tyrosine Kinase↗
PubMed1994

A fifth member of the mammalian G-protein beta-subunit family. Expression in brain and activation of the beta 2 isotype of phospholipase C.

The beta-subunits of the heterotrimeric GTP-binding proteins (G-proteins) are important regulators of G-protein alpha-subunits as well as of certain signal transduction receptors and effectors. We have identified a fifth member of the G-protein beta-subunit family, G beta 5, by molecular cloning. Sequence analysis of recombinant clones revealed an open reading frame of 1,059 base pairs, potentially encoding a protein of 353 amino acids having 53% identity to previously identified G beta-subunit proteins. Northern blot analysis of poly(A)-selected mRNA prepared from different mouse tissues demonstrated that G beta 5 is predominantly expressed in adult brain as two transcripts of 2.3 and 1.75 kilobases. A G beta 5-specific antiserum detected a band of 39.3 kilodaltons on immunoblots of crude membrane proteins isolated from normal mouse brain but not from other tissues. Using a transient cotransfection assay, we showed that a plasmid construct expressing the G beta 5 open reading frame, when coexpressed with different G-protein gamma (G gamma)-subunits, can stimulate the activity of the beta 2 isoform of phosphoinositide-specific phospholipase C (PLC beta 2). The stimulation of PLC beta 2 was most pronounced when G beta 5 was co-expressed with G gamma 2, although activation was observed when G beta 5 was cotransfected with either G gamma 1, G gamma 3, G gamma 5, or G gamma cone. These results prove that G beta 5 can functionally associate with G gamma proteins and is therefore a bona fide member of the G beta protein family. The rather low level of identity between G beta 5 and the other mammalian G beta-subunits may shed light on structural/sequence elements necessary for G beta protein function.

Amino Acid Sequence↗