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Biomedical subjects

J D Scott

Publications and source records attributed to J D Scott.

At least 109 records · Page 6Linked to original sources
PubMed1996

A family with Stickler syndrome type 2 has a mutation in the COL11A1 gene resulting in the substitution of glycine 97 by valine in alpha 1 (XI) collagen.

Stickler syndrome (hereditary arthro-ophthalmopathy) is the commonest inherited cause of retinal detachment and one of the commonest autosomal dominant connective tissue dysplasias. There is clinical and locus heterogeneity with about two thirds of families linked to the gene encoding type II procollagen (COL2A1). Families with Sticklers syndrome type 1 have a characteristic congenital vitreous anomaly and are linked without recombination to markers at the COL2A1 locus. In contrast families with the type 2 variety have a different vitreo-retinal phenotype and are not linked to the COL2A1 gene. Type XI collagen is a quantitatively minor fibrillar collagen related to type V collagen and associated with the more abundant type II collagen fibrils. A mutation in COL11A2, the gene for alpha 2 (XI) procollagen, has recently been found in a family described as having Stickler syndrome, although there was no ocular involvement. Here we show for the first time that a family with the full Type 2 Stickler syndrome including vitreous and retinal abnormalities is linked to the COL11A1 gene and characterise the mutation as a Glycine to Valine substitution at position 97 of the triple helical domain caused by a single base G-->T mutation. These results are the first to provide confirmation that type XI collagen is an important structural component of human vitreous. They also support previous work suggesting that mutations in the genes encoding collagen XI can give rise to some manifestations of Stickler syndrome, but of these, only mutations in COL11A1 will give the full syndrome including the vitreo-retinal features.

Collagen↗
PubMed1996

More on target with protein phosphorylation: conferring specificity by location.

One important regulatory mechanism in the control of phosphorylation events is the subcellular location of phosphatases of kinases. Several serine/threonine phosphatases and kinases have now been shown to be associated with targeting subunits; these contribute to the organization and specificity of signal transduction pathways by favoring the accessibility of their enzymes to certain substrates.

Animals↗
PubMed1996

Early intestinal microparticle uptake in the rat.

This time-course study investigates the early uptake and passage of microparticles across the intestinal mucosa. Single intraoral doses of fluorescent latex particles, 1.82 microns in diameter, were administered to young adult male, nonfasted rats at a dose of 1.88 x 10(9) particles. Peyer's patch regions and mesenteric lymph nodes were collected at 5, 15 and 30 min time points for both bulk tissue and morphological analyses. Particles were found at all experimental time points in macerated intestinal and nodal specimens: particle numbers were higher in proximal than in distal intestine at all time points despite the fact that particle numbers in distal Peyer's patch regions increased with time. Particle numbers in mesenteric lymph nodes also increased with time after administration. Detailed morphological data for several intestinal and nodal tissue compartments showed substantial early uptake of particles by villous epithelium, including goblet cells, but low involvement of follicle-associated cells. The distribution of particles in the lymph nodes confirmed that translocation occurred to all nodal compartments. These studies give confirmatory evidence that uptake and translocation of microparticles may take place as early as 5 min after administration and suggest that intestinal region may influence uptake.

Animals↗
PubMed1996

Systematic expression of the complete coding sequence of apoB-100 does not reveal transmembrane determinants.

We have investigated the hypothesis that apolipoprotein B undergoes a regulated process of translocation into the endoplasmic reticulum (ER) which causes the protein to adopt a transmembrane configuration. Protein segments representing the complete coding sequence of apolipoprotein B were first expressed by in vitro translation of transcripts from seven overlapping transcripts. Two regions were identified (located at residue 2425 and between residues 4149 and 4348) that can undergo incomplete translocation into pancreatic microsomes. Ribosome pausing at these sites uncoupled translation from translocation, leading to the synthesis of large cytoplasmically oriented segments of protein. In contrast, when these two regions were expressed by transfection in cultured cells, transmembrane structures were not detected. Endogenous apolipoprotein B-100 synthesis in HepG2 cells generates a spectrum of nascent chains, indicating that ribosome pausing can also occur in intact cells. However, the cellular pause products were cotranslationally translocated. While endogenous apolipoprotein B-100 in HepG2 cells was fully translocated, discrete proteolytic fragments were generated from the amino terminus of the protein when proteases gained access to the lumen of permeabilized microsomes. These products were similar in size and sequence to apoliprotein B proteolytic fragments previously ascribed as the luminal domains of transmembrane apoB-100 molecules (Du, E. Z., Kurth, J., Wang, S. L., Humiston, P., and Davis, R. A. 1994. J. Biol. Chem. 269: 24169-24176).

Amino Acid Sequence↗
PubMed1995

Cloning and characterization of A-kinase anchor protein 100 (AKAP100). A protein that targets A-kinase to the sarcoplasmic reticulum.

Differential localization of the type II cAMP-dependent protein kinase (PKA) is achieved by interaction of the regulatory subunit (RII) with A-kinase anchor proteins (AKAPs). Anchoring is a likely means to adapt PKA for regulation of cAMP-responsive events through colocalization of the kinase with preferred substrates. Using an interaction cloning strategy with an RII alpha protein probe, we have identified a 655-amino acid protein (named AKAP100). Recombinant AKAP100, expressed in Escherichia coli, binds RII alpha in a solid-phase overlay assay. The cellular and subcellular distribution of AKAP100 was analyzed by various methods. Northern blot analysis with the AKAP100 cDNA as a probe detected an 8-kilobase message in some human tissues including various brain regions; however, the message was predominately expressed in cardiac and skeletal muscle. Anti-AKAP100 antibodies confirmed expression in the rat cardiac and skeletal muscle cell lines, H9c2 and L6P, whereas immunohistochemical analysis revealed that AKAP100 was localized to the sarcoplasmic reticulum of both cell types. RII was also detected in these regions. AKAP100 was detected in preparations of RII purified from L6P cell extracts by cAMP-agarose affinity chromatography. Collectively, these results suggest that AKAP100 functions to maintain the type II PKA at the sarcoplasmic reticulum.

A Kinase Anchor Proteins↗
PubMed1995

Association of protein kinase A and protein phosphatase 2B with a common anchoring protein.

Specificity of protein kinases and phosphatases may be achieved through compartmentalization with preferred substrates. In neurons, adenosine 3', 5'-monophosphate (cAMP)-dependent protein kinase (PKA) is localized at postsynaptic densities by association of its regulatory subunit with an A kinase anchor protein, AKAP79. Interaction cloning experiments demonstrated that AKAP79 also binds protein phosphatase 2B, or calcineurin (CaN). A ternary complex of PKA, AKAP, and CaN was isolated from bovine brain, and colocalization of the kinase and the phosphatase was established in neurites of cultured hippocampal neurons. The putative CaN-binding domain of AKAP79 is similar to that of the immunophilin FKBP-12, and AKAP79 inhibited CaN phosphatase activity. These results suggest that both PKA and CaN are targeted to subcellular sites by association with a common anchor protein and thereby regulate the phosphorylation state of key neuronal substrates.

A Kinase Anchor Proteins↗
PubMed1995

P-31 changes as a measure of therapy response in resistant and sensitive osteosarcomas implanted into nude mice.

OBJECTIVE: To determine if changes in PCr/Pi and PME can be used to predict lack of tumor response to chemotherapy in a murine model of a chemotherapy-resistant human osteosarcoma. MATERIAL AND METHODS: Cisplatin-resistant sublines were grown from high-grade cisplatin-sensitive human osteosarcoma. Surface coil localized 31P NMR spectroscopy of implanted cisplatin-resistant and sensitive osteosarcoma tumors in nude mice was performed. RESULTS: A cisplatin-resistant subline of a sensitive human osteosarcoma was developed that was five times more resistant to cisplatin than the parent cell line. Our NMR data shows a statistically significant difference in the change in the PCr/Pi ratio after treatment between sensitive and resistant osteosarcomas at the alpha = 0.05 level. Changes in PME were seen in the sensitive tumors but were not statistically significant. CONCLUSIONS: Changes in PCr/Pi predict lack of tumor treatment response in human osteosarcoma implanted into nude mice with a specificity of 70% and a sensitivity of 54%. Monitoring of PCr/Pi in human osteosarcoma patients may allow detection of response to chemotherapy before conventional imaging techniques.

Animals↗
PubMed1995

Adverse events associated with trimethoprim-sulfamethoxazole and atovaquone during the treatment of AIDS-related Pneumocystis carinii pneumonia.

Atovaquone was compared to trimethoprim-sulfamethoxazole (TMP-SMZ) for the relationship of time receiving therapy, plasma drug concentrations, and incidence of adverse reactions in patients with AIDS-associated Pneumocystis carinii pneumonia. Treatment-limiting adverse events occurred in 9% of atovaquone-treated patients and 24% of TMP-SMZ-treated patients. Adverse events usually did not occur before day 7 for either treatment. Only the incidence of rash increased with increasing plasma concentrations of atovaquone. The incidence of anemia, neutropenia, and azotemia increased with increasing trimethoprim plasma concentration, while other adverse events (gastrointestinal disorders, rash, fever, and liver function abnormalities) were independent of plasma drug concentration.

AIDS-Related Opportunistic Infections↗
PubMed1994

Type II regulatory subunit (RII) of the cAMP-dependent protein kinase interaction with A-kinase anchor proteins requires isoleucines 3 and 5.

Compartmentalization of the type II cAMP-dependent protein kinase is maintained by association of the regulatory subunit (RII) with A-Kinase Anchor Proteins (AKAPs). In previous studies (Scott, J. D., Stofko, R. E., McDonald, J. R., Comer, J. D., Vitalis, E. A., and Mangili J. (1990) J. Biol. Chem. 265, 21561-21566) we have shown that dimerization of RII alpha was required for interaction with the cytoskeletal component microtubule-associated protein 2. In this report we show that the localization and dimerization domains of RII alpha are contained within the first thirty residues of each RII protomer. RII des-5 (an amino-terminal deletion mutant lacking residues 1-5) was unable to bind AKAPs but retained the ability to dimerize. RII alpha I3A,I5A (a mutant where isoleucines 3 and 5 were replaced with alanine) was unable to bind a variety of AKAPs. Mutation of both isoleucines decreased AKAP binding without affecting dimerization, cAMP binding, or the overall secondary structure of the protein. Measurement of RII alpha I3A,I5A interaction with the human thyroid AKAP, Ht 31, by two independent methods suggests that mutation of isoleucines 3 and 5 decreases affinity by at least 6-fold. Therefore, we propose that two isoleucine side chains on each RII protomer are principle sites of contact with the conserved amphipathic helix binding domain on AKAPs.

Amino Acid Sequence↗