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J R Bourne

Publications and source records attributed to J R Bourne.

17 recordsLinked to original sources
PubMed1992

GTP-binding mutants of rab1 and rab2 are potent inhibitors of vesicular transport from the endoplasmic reticulum to the Golgi complex.

We have examined the role of ras-related rab proteins in transport from the ER to the Golgi complex in vivo using a vaccinia recombinant T7 RNA polymerase virus to express site-directed rab mutants. These mutations are within highly conserved domains involved in guanine nucleotide binding and hydrolysis found in ras and all members of the ras superfamily. Substitutions in the GTP-binding domains of rab1a and rab1b (equivalent to the ras 17N and 116I mutants) resulted in proteins which were potent trans dominant inhibitors of vesicular stomatitis virus glycoprotein (VSV-G protein) transport between the ER and cis Golgi complex. Immunofluorescence analysis indicated that expression of rab1b121I prevented delivery of VSV-G protein to the Golgi stack, which resulted in VSV-G protein accumulation in pre-Golgi punctate structures. Mutants in guanine nucleotide exchange or hydrolysis of the rab2 protein were also strong trans dominant transport inhibitors. Analogous mutations in rab3a, rab5, rab6, and H-ras did not inhibit processing of VSV-G to the complex, sialic acid containing form diagnostic of transport to the trans Golgi compartment. We suggest that at least three members of the rab family (rab1a, rab1b, and rab2) use GTP hydrolysis to regulate components of the transport machinery involved in vesicle traffic between early compartments of the secretory pathway.

Amino Acid Sequence↗
PubMed1991

Isoprenoid modification of rab proteins terminating in CC or CXC motifs.

Mevalonate starvation of hamster fibroblasts resulted in a shift of rab1b from the membrane to the cytosolic fraction, suggesting that rab1b depends upon an isoprenoid modification for its membrane localization. rab1b and rab3a proteins expressed in insect cells incorporated a product of [3H]mevalonate, and gas chromatography analysis of material released by Raney nickel cleavage demonstrated that rab1b and rab3a are modified by geranylgeranyl groups. Additionally, in vitro prenylation analysis demonstrated farnesyl modification of H-ras but geranylgeranyl modification of five rab proteins (1a, 1b, 2, 3a, and 6). Together, these results suggest that the carboxyl-terminal CC/CXC motifs (X = any amino acid) specifically signal for addition of geranylgeranyl, but not farnesyl, groups. A rab1b mutant protein lacking the two carboxyl-terminal cysteine residues was not prenylated in vitro. However, since a mutant H-ras protein that terminates with tandem cysteine residues was also not modified, the CC motif may be essential, but not sufficient, to signal prenylation of rab1b. Finally, rab1b and rab3a proteins were not efficient substrates for either farnesyl- or geranylgeranyltransferase activities that modify CAAX-containing proteins (A = any aliphatic amino acid). Therefore, rab proteins may be modified by a prenyltransferase(s) distinct from the prenyltransferases that modify carboxyl-terminal CAAX proteins.

Amino Acid Sequence↗
PubMed1991

Rab1b regulates vesicular transport between the endoplasmic reticulum and successive Golgi compartments.

We report an essential role for the ras-related small GTP-binding protein rab1b in vesicular transport in mammalian cells. mAbs detect rab1b in both the ER and Golgi compartments. Using an assay which reconstitutes transport between the ER and the cis-Golgi compartment, we find that rab1b is required during an initial step in export of protein from the ER. In addition, it is also required for transport of protein between successive cis- and medial-Golgi compartments. We suggest that rab1b may provide a common link between upstream and downstream components of the vesicular fission and fusion machinery functioning in early compartments of the secretory pathway.

Animals↗
PubMed1990

Farnesol modification of Kirsten-ras exon 4B protein is essential for transformation.

Oncogenic forms of ras proteins are synthesized in the cytosol and must become membrane associated to cause malignant transformation. Palmitic acid and an isoprenoid (farnesol) intermediate in cholesterol biosynthesis are attached to separate cysteine residues near the C termini of H-ras, N-ras, and Kirsten-ras (K-ras) exon 4A-encoded proteins. These lipid modifications have been suggested to promote or stabilize the association of ras proteins with membranes. Because preventing isoprenylation also prevents palmitoylation, examining the importance of isoprenylation alone has not been possible. However, the oncogenic human [Val12]K-ras 4B protein is not palmitoylated but is isoprenylated, membrane associated, and fully transforming. We therefore constructed mutant [Val12]K-ras 4B proteins that were not isoprenylated to examine the effects of isoprenylation in the absence of palmitoylation. The nonisoprenylated mutant proteins both failed to associate with membranes and did not transform NIH 3T3 cells. In addition, inhibition of isoprenoid and cholesterol synthesis with the drug compactin also decreased [Val12]K-ras 4B protein isoprenylation and membrane association. These results unequivocally demonstrate that isoprenylation, rather than palmitoylation, is essential for ras membrane binding and ras transforming activity. These findings clearly indicate the biological significance of ras protein modification by farnesol and suggest that this modification may be important for facilitating the processing, trafficking, and biological activity of other isoprenylated proteins. Because K-ras is the most frequently activated oncogene in a wide spectrum of human malignancies, study of this pathway could lead to important therapeutic treatments.

Animals↗
PubMed1979

Quantitative indices of clinical uremia.

The disabling, dialysis-responsive symptoms of clinical uremia primarily represent impaired functions of the nervous system. Accordingly, these studies used several quantitative electrophysiologic and cognition-dependent probes of nervous system function: peripheral nerve-conduction velocity, response latency and amplitude; electroencephalographic (EEG) power-spectrum analysis; visual evoked response latency; EEG responses to photic driving; and measures of integrated behavioral performance that stressed sustained and selective attention, reaction time, speed of decision-making, short-term memory, and mental manipulation of symbols. Probes of quanitified central nervous system function consistently revealed impairments that varied directly with the degree of renal failure, that improved following onset of maintenance dialysis, and that improved still further after successful renal transplantation. In contrast, measures of peripheral nerve function were generally unchanged. Neurobehavioral measures that are relevant to uremic symptomatology provide quantitative estimates of the clinically significant, whole-organism biologic effects of renal failure and its several treatments. Applications of such measures in studies of symptomatic states other than uremia are indicated also.

Adolescent↗
PubMed1978

Visually evoked cortical potentials in renal failure: transient potentials.

Transient visually evoked cortical potentials (VECPs) were recorded from patients with renal disease. Changes in VECP latencies are described for undialyzed patients, patients receiving dialysis therapy, and patients who received kidney transplants. Characteristics of VECP latencies in these patient groups as well as examples of changes in latency values with respect to time for two individual patients are examined. The basic overall finding is that the VECP latencies increase as a patient's clinical condition deteriorates and normalize as the condition improves.

Adolescent↗
PubMed1978

Quantitative assessment of photic driving in renal failure.

Photically driven EEGs were recorded from patients with renal disease using photic stimulation at integer rates between 3 and 12 flashes/sec. Changes in the structure of the power spectrum of the potentials produced by this stimulus paradigm are described as a function of a patient's clinical state. The basic overall findings are that (1) harmonic activity is attenuated while activity below the fundamental driving frequency is increased as a patient's clinical condition deteriorates, and (2) that these effects are substantially reversed and controlled by means of dialysis and renal transplantation.

Adolescent↗
PubMed1975

Quantitative assessment of the electroencephalogram in renal disease.

EEGs wre recorded from renal patients to determine if there are quantifiable characteristic changes in the EEG was quantified by calculating the percentage of spectral power in the bandwidth 3-7 c/sec referrred to a frequency range of 3-13 c/sec and by computing the mean frequency of the dominant rhythm in the EEG. Blood urea nitrogen and creatinine concentrations, as well as a self-assessment of the patient's clinical condition, were recorded. The general finding of this research is that EEG slowing, as evaluated by power spectral techniques, is correlated with uremia-associated variables. 1. In a non-dialyzed patient population with renal failure, slowing in the EEG was found to be directly corelated with increased creatinine concentrations. 2. Quantitative measures of slow wave activity computed using power spectral techniques were found to be highly corelated with an estimate of slowing made by an electroencephalographer. 3. Compared with undialyzed azotemic patients, malignant hypertensive patients with comparable serum creatinine concentrations typically displayed increased slow wave activity, while slowing was generally reduced in the dialyzed patient population. 4. A series of EEGs recorded from one patient during the first three dialyses of her life revealed that slow wave activity decreased during each successive dialysis. In another patient, all quantified EEG values recorded prior to renal transplantation significantly improved after transplantation...

Adolescent↗
PubMed1975

Neurotoxicity in uremia.

The dialysis patient is aware that his behavior evokes reciprocal and complicating responses from important people in his environment. These interactions are perceived and conducted by neurochemical mechanisms which may be impaired in the abnormal chemical environment imposed in renal failure. It is the behaviors we comprehend as indicators of disordered nervous mechanisms. Therefore, it is logical that neurophysiological and neurobehavioral phenomena should be measured quantitatively in order 1) to estimate objectively the patients' success in achieving the goal of maintenance dialysis treatment, 2) to assess the comparative adequacy of dialysis regimens and 3) to provide objective endpoint measures which are relevant to uremia for further investigations of the etiology and pathogenesis of these critically significant uremic manifestations. Our experimental results illustrate that: 1) measures of conduction velocity, distal latency and response amplitudes, as employed by us, were relatively insensitive in the patients and circumstances studied; 2) several neurophysiological measures, i.e., the spontaneous EEG, VER latency and, perhaps, photic stimulation, on the other hand, are highly correlated with the severity of renal failure; 3) behavioral measures of sustained attention and alertness (TMT), of short-term memory (ASTM) and of cognitive manipulation of symbols (AR) are also highly correlated with the severity of renal failure; 4) some measured abnormalities improve following dialysis, but not always to normal--three residual impairments may indicate dialysis in adequacy; 5) several of these measures can provide objective evidence for adequacy of dialysis and other clinical and treatment effects in patients with renal failure.

Auditory Cortex↗