Structural analysis of the MAP kinase ERK2 and studies of MAP kinase regulatory pathways.
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Amino Acid Sequence↗
Biomedical subjects
D Robbins
Publications and source records attributed to D Robbins.
Apolipoprotein E polymorphism and LDL size in a biethnic population.
Polymorphisms in the apolipoprotein E (apoE) phenotype (especially E4) are associated with increased cardiovascular risk and particularly with increased concentrations of LDL cholesterol. Little is known, however, about whether alterations in LDL size are associated with the apoE4 phenotype. LDL size was determined by gradient gel electrophoresis, and apoE phenotype was determined by isoelectric focus in 337 nondiabetic subjects from the San Antonio Heart Study, a population-based study of diabetes and cardiovascular risk factors in Mexican Americans and non-Hispanic whites. ApoE4 was associated not only with increases in LDL cholesterol concentrations but also with decreased LDL size. After adjustment for age, sex, body mass index, waist-to-hip ratio, triglyceride, HDL cholesterol, fasting insulin, and diabetic status, the apoE phenotype remained significantly related to LDL size (A) in both men (apoE23, 260.0; apoE33; 256.3; and apoE34, 252.6; P = .01) and women (apoE23, 261.7; apoE33, 257.9; and apoE34, 256.7, P = .045). Variations in apoE phenotype are associated not only with changes in the absolute concentration of LDL cholesterol but also with changes in its composition. These changes are only partly explained by associations of apoE with insulin, triglyceride, and HDL cholesterol.
Adult↗
MEKK1 phosphorylates MEK1 and MEK2 but does not cause activation of mitogen-activated protein kinase.
A constitutively active fragment of rat MEK kinase 1 (MEKK1) consisting of only its catalytic domain (MEKK-C) expressed in bacteria quantitatively activates recombinant mitogen-activated protein (MAP) kinase/extracellular signal-regulated protein kinase (ERK) kinases 1 and 2 (MEK1 and MEK2) in vitro. Activation of MEK1 by MEKK-C is accompanied by phosphorylation of S218 and S222, which are also phosphorylated by the protein kinases c-Mos and Raf-1. MEKK1 has been implicated in regulation of a parallel but distinct cascade that leads to phosphorylation of N-terminal sites on c-Jun; thus, its role in the MAP kinase pathway has been questioned. However, in addition to its capacity to phosphorylate MEK1 in vitro, MEKK-C interacts with MEK1 in the two-hybrid system, and expression of mouse MEKK1 or MEKK-C in mammalian cells causes constitutive activation of both MEK1 and MEK2. Neither cotransfected nor endogenous ERK2 is highly activated by MEKK1 compared to its stimulation by epidermal growth factor in spite of significant activation of endogenous MEK. Thus, other as yet undefined mechanisms may be involved in determining information flow through the MAP kinase and related pathways.
Animals↗
Shk1, a homolog of the Saccharomyces cerevisiae Ste20 and mammalian p65PAK protein kinases, is a component of a Ras/Cdc42 signaling module in the fission yeast Schizosaccharomyces pombe.
We describe a protein kinase, Shk1, from the fission yeast Schizosaccharomyces pombe, which is structurally related to the Saccharomyces cerevisiae Ste20 and mammalian p65PAK protein kinases. We provide genetic evidence for physical and functional interaction between Shk1 and the Cdc42 GTP-binding protein required for normal cell morphology and mating in S. pombe. We further show that expression of the STE20 gene complements the shk1 null mutation and that Shk1 is capable of signaling to the pheromone-responsive mitogen-activated protein kinase cascade in S. cerevisiae. Our results lead us to propose that signaling modules composed of small GTP-binding proteins and protein kinases related to Shk1, Ste20, and p65PAK, are highly conserved in evolution and participate in both cytoskeletal functions and mitogen-activated protein kinase signaling pathways.
Amino Acid Sequence↗
A preponderance of small dense LDL is associated with specific insulin, proinsulin and the components of the insulin resistance syndrome in non-diabetic subjects.
Recently, the presence of small dense low density lipoprotein (LDL) has been postulated to be a stronger risk factor for coronary heart disease than large LDL. While small dense LDL has been associated with individual components of the insulin resistance syndrome such as hypertension, high triglyceride level, low high density (HDL) cholesterol, and diabetes mellitus, there has been little work exploring whether LDL size is decreased in subjects with multiple metabolic disorders. We examined the association of LDL size and pattern to specific insulin (which does not cross-react with proinsulin), proinsulin, increased triglyceride, decreased HDL, hypertension and impaired glucose tolerance in 488 non-diabetic subjects from the San Antonio Heart Study. LDL size was significantly related to specific insulin, proinsulin and the fasting proinsulin/insulin ratio. Small dense LDL was significantly associated with high triglyceride level, decreased HDL cholesterol, hypertension and impaired glucose tolerance. LDL size (A) decreased in a stepwise fashion with increasing number of the metabolic disorders described above (zero 262.6 +/- 9.4; one 257.0 +/- 9.3; two 256.4 +/- 9.4; three 249.0 +/- 9.1; and four 244.9 +/- 9.0). These results were similar in men and women and in non-Hispanic whites and Mexican Americans. The association between LDL size and the number of metabolic disorders remained statistically significant even after adjustment for obesity, body fat distribution, gender, ethnicity, proinsulin and insulin concentrations. Furthermore, decreases in LDL size are also significantly associated with both a selective beta-cell defect (as estimated by the fasting proinsulin/insulin ratio) and insulin resistance (as estimated by the fasting insulin concentrations) although the association was somewhat stronger for the latter. We conclude that small dense LDL may form part of the insulin resistance syndrome in non-diabetic subjects.
Diabetes Mellitus↗
Reduction of glycosylated hemoglobin and postprandial hyperglycemia by acarbose in patients with NIDDM. A placebo-controlled dose-comparison study.
OBJECTIVE: To compare the safety and efficacy of three doses of acarbose (100, 200, and 300 mg three times daily) with placebo for the treatment of non-insulin-dependent diabetes mellitus (NIDDM) in patients maintained on dietary therapy alone. RESEARCH DESIGN AND METHODS: This multicenter double-blind placebo-controlled trial was 22 weeks in duration. The trial consisted of a 2-week screening period, a 4-week placebo run-in period, and a 16-week double-blind treatment period. The primary measure of drug efficacy was the mean change from baseline in HbA1c levels. Additional efficacy variables included the mean change from baseline in fasting and postprandial plasma glucose and serum insulin levels. RESULTS: After 16 weeks of treatment, acarbose-treated patients had statistically significant reductions in mean HbA1c levels of 0.78, 0.73, and 1.10% (relative to placebo) in the 100-, 200-, and 300-mg t.i.d. groups, respectively. Significant reductions in fasting and postprandial plasma glucose levels, glucose area under the time-concentration curve, and maximum glucose concentration were also observed in acarbose-treated patients. Although there were no statistically significant differences among the 100-, 200-, and 300-mg treatment groups, there was a trend toward a dose-response relationship for most plasma glucose variables that were measured. Gastrointestinal side effects (e.g., abdominal pain, flatulence, and diarrhea) and serum transaminase elevations (e.g., aspartate aminotransferase [AST] and alanine aminotransferase [ALT] were more frequently reported in the acarbose-treated patients than in the placebo-treated control patients. Transaminase elevations occurred only at the 200-, and 300-mg dosages and were readily reversible on discontinuation of treatment. CONCLUSIONS: Acarbose at doses of 100, 200, and 300 mg administered three times daily for 16 weeks significantly reduced HbA1c levels and postprandial hyperglycemia. Treatment with acarbose is a safe and effective adjunct to dietary therapy for the treatment of NIDDM.
Acarbose↗
A subgroup of human VH3 germline genes that encode a high-avidity synovial rheumatoid factor.
We have previously derived and identified a highly avid monoclonal IgM rheumatoid factor (mRF), C6, from unstimulated rheumatoid synovial cells (RSC). At the time, the closet VH germline gene, VH26, demonstrated only 88% homology with C6. To identify the germline counterpart of C6, genomic DNA from the same rheumatoid arthritis (RA) patient from whom C6 was derived was used in the polymerase chain reaction (PCR). Four of the six closely related germline genes that we sequenced had exonic regions that were identical with the VH region of C6 cDNA. These six germline sequences differed in their intronic regions, suggesting that they were distinct, but closely related genomic sequences. To further evaluate the extent of these related genes we identified nine additional germline genes having VH-encoding exons that were 86-97% identical to the C6 cDNA sequence. Furthermore, we examined the polymorphic nature of the C6 VH gene using single strand conformation polymorphism (SSCP), and identified two peaks, confirming the existence of highly homologous genes. The sequence and polymorphism data suggest that: (1) the VH region of the high avidity mRF C6 was derived from an unmutated germline gene; (2) C6 was encoded by a VH gene belonging to a set of homologous genes within the larger VH3 family; and (3) in addition to somatic rearrangements of B-cell genes and antigen-driven somatic mutation, gene duplication and conversion events of germline genes could be important in generating diversity and polyclonality among high-affinity pathogenic autoantibodies.
Amino Acid Sequence↗
Atomic structure of the MAP kinase ERK2 at 2.3 A resolution.
The structure of the MAP kinase ERK2, a ubiquitous protein kinase target for regulation by Ras and Raf, has been solved in its unphosphorylated low-activity conformation to a resolution of 2.3 A. The two domains of unphosphorylated ERK2 are farther apart than in the active conformation of cAMP-dependent protein kinase and the peptide-binding site is blocked by tyrosine 185, one of the two residues that are phosphorylated in the active enzyme. Activation of ERK2 is thus likely to involve both global and local conformational changes.
Adenosine Triphosphate↗
Factors associated with dangerous behavior in forensic inpatients: results from a pilot study.
This study was designed to identify risk factors associated with violence within a forensic inpatient hospital setting. The primary purpose was to develop a screening tool to aid in the rapid identification of patients requiring high versus low security ward placement. Subjects included 232 consecutive admissions during a five-month period to a 300-bed forensic division within a public-sector psychiatric hospital. Demographic, historical, and current clinical variables were collected, and dangerous behaviors were documented by nursing staff on a daily basis. The associations between dangerous outcomes and various risk factors were examined using chi-square or t tests, as appropriate. Stepwise logistic regression analysis was performed to assess the contribution of each risk factor significantly associated with violent behavior. A strong association was found between 10 risk factors and the incidence of violence in our forensic population. These factors included current clinical factors (grossly inappropriate behavior observed on the ward, an assessment of current escape risk); historical factors (psychiatric hospitalization history, criminal history variables, and a self-report of prior violence); and one demographic variable (length of stay). Of the 10 risk factors then included in the stepwise regression analysis, four contributed significantly to the final predictive model. Grossly inappropriate behavior, patient self-report of prior violence, history of psychiatric hospitalization, and a history of 10 or more prior psychiatric hospitalizations were most predictive of a violent outcome. Although situational and environmental variables also must be incorporated into final assessments and decisions, the identified significant variables can provide an additional tool in the rapid assessment of violence potential.
Dangerous Behavior↗
The mitogen-activated protein kinases, ERK1 and ERK2.
The MAP kinases are ubiquitous enzymes that are activated in a complex fashion and inactivated by multiple phosphatases including a dedicated dual specificity enzyme. These kinases have a diverse array of substrates with important functions that result in their substantial regulatory impact. The ERK/MAP kinase cascade displays not only downstream but also upstream interactions as well as cross talk with other signaling pathways which fine tunes the cascade in a cell type-specific fashion. Transforming agents utilize this cascade in inducing cell proliferation.
Animals↗
The interaction of SV40 small tumor antigen with protein phosphatase 2A stimulates the map kinase pathway and induces cell proliferation.
Interaction with SV40 small tumor antigen (small t) compromised the ability of multimeric protein phosphatase 2A to inactivate the mitogen-activated protein kinase ERK1 and the mitogen-activated protein kinase kinase MEK1. Transient expression of small t in CV-1 cells activated MEK and ERK but did not affect Raf activity. Small t stimulated the growth of quiescent CV-1 cells almost as effectively as did serum. Coexpression of kinase-deficient ERK2 blocked most, but not all, of the proliferation caused by small t. Activation of the mitogen-activated protein kinase pathway and stimulation of cell growth were dependent on the interaction of small t with protein phosphatase 2A. These findings indicate that SV40 small t is capable of inducing cell growth through blockade of protein phosphatase and deregulation of the mitogen-activated protein kinase cascade.
Animals↗
Monoclonal IgM rheumatoid factors generated from synovial B cells of rheumatoid arthritis patients react with beta 2-microglobulin. Monoclonal RF react with beta 2m.
Four of 15 monoclonal human IgM rheumatoid factors (RF) derived from synovial B cells of patients with rheumatoid arthritis showed positive ELISA reactions with human beta 2-microglobulin. These findings were different from those previously noted using IgM RF derived from monoclonal Waldenstrom's paraproteins or the IgM components of mixed cryoglobulins, and resembled the anti-beta 2 microglobulin specificity of polyclonal IgM RF from patients with rheumatoid arthritis. Reactions of monoclonal IgM synovial RF with overlapping 7-mers of beta 2m sequence indicated major regions of positive reactivity at positions 57-64 and 89-95 which were maintained in the presence of high salt (300 mM NaCl) conditions. Glycine substitution of each residue within RF-reactive beta 2m regions indicated that tryptophanes at position 60 and 95, lysine at 58, phenylalanine at 62, valine at 93 and arginine at 97 constituted important single amino acids for the reactive epitopes. These findings indicate that clonally restricted human IgM RF derived from diseased tissues of patients with RA show anti-beta 2m reactivity similar to polyclonal RF from the same patients. This particular fine specificity is not present in monoclonal RF derived from patients with Waldenstrom's or mixed cryoglobulins showing anti-gamma-globulin activity.
Amino Acid Sequence↗
Monitoring of antiviral therapy with quantitative evaluation of HBeAg: a comparison with HBV DNA testing.
The serological endpoint of response in the treatment of chronic hepatitis B is the loss of hepatitis B virus DNA and HBeAg. Because the quantitative measurement of hepatitis B virus DNA in serum has been shown to be useful for monitoring and predicting response to interferon-alpha therapy, we decided to evaluate whether changes in HBeAg concentration could also be used in this manner. Twenty-nine patients who were initially positive for HBeAg and HBV DNA were serially evaluated for HBeAg concentration with a microparticle-capture enzyme immunoassay. HBeAg levels in serum were calculated by means of comparison with a standard curve of fluorescence rate vs. HBeAg concentration. The results, expressed in milliunits per milliliter, were compared with hepatitis B virus DNA levels determined by means of solution hybridization. The baseline HBeAg concentration proved to be the best independent predictor of response on stepwise Cox regression analysis (p = 0.026). Similar disappearance curves were observed for the two markers, although hepatitis B virus DNA became undetectable at an earlier interval in 13 of 16 cases (81%). In the 16 responders, a decline in HBeAg concentration of more than 90% was observed by wk 12 of therapy (mean +/- S.D., 95% +/- 13%). Nonresponders did not demonstrate such steep declines in HBeAg values by wk 12 (mean +/- S.D., 45% +/- 27%), and levels tended to increase at subsequent time points. We conclude that serial monitoring of HBeAg concentration with a technique that should be readily adaptable to clinical laboratories may be useful in the initial evaluation and monitoring of patients undergoing antiviral therapy.
DNA, Viral↗
Human monoclonal rheumatoid factor derived from rheumatoid synovial cells monospecific for rabbit IgG.
Rheumatoid factors (RF) in rheumatoid arthritis (RA) are polyclonal autoantibodies directed against antigenic epitopes located in the Fc portion of the IgG molecule. Hybridoma technology has overcome the difficulty of their polyclonality, so that monoclonal RF (mRF) can be examined for their individual binding specificities and genetics. We isolated a monoclonal IgM RF secreting hybridoma (designated H4) from the rheumatoid synovial cells (RSC) of a patient with RA. H4 bound specifically with rabbit IgG (RIgG) and had no human IgG (HIgG) reactivity. By direct binding ELISA and absorption experiments, 6% of the RIgG reactive RSC RF in this patient with RA was monospecific for RIgG. H4 was tested against RIgG F(ab')2 and pFc' fragments, and bound only to the pFc' fragment (CH3 domain). Moreover, H4 mRF had high avidity for RIgG in a capture ELISA. Total RNA was extracted and the variable region heavy (VH) and light (VL) chain cDNA were amplified using polymerase chain reaction technology. Sequence analysis of the IgM RF VH and VL chain genes indicated usage of the VH26 germline gene (VhIII gene family) and a new V lambda germline gene. Our results suggest preferential use of restricted germline genes in the formation of autoantibodies in human autoimmune diseases. The pathological significance of RIgG specific RF is still unclear. However, this finding suggests that all RSC RF production may not necessarily be induced by autologous IgG.
Amino Acid Sequence↗
Allotype-dependent stimulation of peripheral blood and synovial lymphocytes by IgG3 in rheumatoid arthritis.
The immunopathologic process of rheumatoid arthritis (RA) is primarily expressed in the synovium where rheumatoid factor (RF) synthesis is concentrated. We hypothesized that RF synthesized by rheumatoid synovial cells (RSC) may be driven via a T cell-mediated immune response developed against IgG3 epitopes. To identify and characterize specific RSC RF epitopes and T cell antigens, two 28 amino acid peptides homologous with the C-terminus of IgG1 (P1) and IgG3 [G3m(5)] (P3) were synthesized and used in RF-binding studies and lymphocyte proliferation assays. Our results indicate that (i) the C-terminus of the CH3 domain contains epitopes for IgG3-reactive RSC RF; (ii) IgG3-reactive RSC RF binds primarily to IgG3 [G3m(5)]; (iii) P3 stimulated proliferation of T lymphocytes from both RA peripheral blood and RSC; and (iv) RF production was enhanced by P3 in selected RA cell cultures. These observations suggest that the C-terminus of IgG3 allotype G3m(5) may be important in T cell activation and RF production in RA.
Amino Acid Sequence↗
Prenatal detection and management of quadruple nuchal cord. A case report.
As imaging techniques improve, nuchal cords are often being identified prior to labor. Nuchal cords are encountered in up to 25% of deliveries and are usually not clinically significant. They may, however, have devastating results. Nuchal cords may be associated with abnormal fetal activity and testing, but there is often no clinical warning of a nuchal cord during pregnancy, labor or delivery. We report a case of a quadruple nuchal cord discovered after the mother complained of decreased fetal movement. The clinical course of this patient highlights many of the difficulties encountered in the management of antenatally diagnosed nuchal cords, particularly in the preterm fetus.
Acidosis↗
Home visits for pregnant diabetic women: environmental assessment as a basis for nursing intervention.
A major goal in management of pregnancies complicated with diabetes is strict control of blood glucose levels. After glucose control is achieved, home visits by the Perinatal Clinical Nurse Specialist (PCNS) who has advanced knowledge and skills in the management of high risk pregnancies can be invaluable. The PCNS systematically assesses the client's environment and its effects upon the client's ability to carry out the many therapeutic self-care demands associated with the diabetic pregnancy. These include the client's ability to: identify and act upon episodes of hypo- or hyperglycemia, monitor blood glucose levels, self-administer insulin, self-monitor fetal activity, and identify and act upon danger signs in pregnancy and signs of labor. The PCNS, by assessing and planning within the framework of the client's environment, facilitates continued control of diabetes as the woman makes the transition from the hospital to the home and/or work environment.
Female↗
An immunological study in patients with seborrhoeic dermatitis.
The humoral and cellular immune-status was studied in 30 patients with seborrhoeic dermatitis. Increased frequencies of natural killer cells were found in 46% of patients. Furthermore, subnormal mitogen stimulation responses were demonstrated in 13 patients, whereas two individuals were found to have very high numbers of activated T lymphocytes in peripheral blood. Higher-than-normal total serum IgG and IgA was observed in 14 and 11 patients, respectively. For nine of 12 patients with skin lesions, dermal perivascular cell infiltrates were seen. The majority of the infiltrating cells reacted with anti-CD4 antibodies. HLA-DR-expressing keratinocytes were found in two biopsies. The study suggests that patients with seborrhoeic dermatitis may have depressed T-cell function. This could have a bearing on their susceptibility to the Pityrosporum ovale-associated dermatitis. The very high frequencies of activated T cells observed in the peripheral blood of two otherwise healthy seborrhoeic individuals suggests that intermittent systemic immune activation may occur. Seborrhoeic dermatitis is a common skin disease. It can be diagnosed by its characteristic red to yellow-brown lesions covered with greasy scales distributed in areas with a high number of sebaceous glands, such as the scalp, face and upper trunk. There is an association between seborrhoeic dermatitis and the lipophilic yeast Pityrosporum ovale but its exact aetiological role is not known. The yeast is a member of the normal cutaneous flora but also the aetiological agent of pityriasis versicolor and Pityrosporum folliculitis. P. ovale can activate complement via the direct and alternative pathways. This may play some part in the induction of inflammation.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗