Biomedical subjects

J M Roberts

Publications and source records attributed to J M Roberts.

At least 55 records · Page 3Linked to original sources

PubMed2000

Invasive cytotrophoblasts manifest evidence of oxidative stress in preeclampsia.

In preeclampsia, poor placental perfusion may result in maternal endothelial dysfunction, but the pathways involved are largely unknown. Candidate placental mediators include products of oxidative stress released into the maternal circulation. Xanthine oxidase has been implicated in postischemic-reperfusion injury via the generation of superoxide anion radicals (superoxide; O(2)(.-)) and hydrogen peroxide. We examined placentas and placental bed curettings and/or biopsies from preeclamptic control pregnant women to test the hypothesis that xanthine oxidase is a mediator of oxidative stress in placentas from women with preeclampsia. The expression of xanthine dehydrogenase/xanthine oxidase holoenzyme and the activity of xanthine oxidase, the isoform known to generate reactive oxygen species, were increased in a subpopulation of cytotrophoblasts of preeclamptic women. Additionally, the expression of superoxide dismutase, which would scavenge superoxide produced by xanthine oxidase, was reduced in the same cells. Furthermore, fluorescence immunostaining for nitrotyrosine, which was suggestive of superoxide-nitric oxide interactions to form peroxynitrite anion (ONOO(-)) in vivo, was increased in these cells and in villous vessels. Thus, our data indicate an increased capacity of placental cells to generate reactive oxygen species in preeclampsia.

Adult↗

PubMed2000

Preeclampsia: what we know and what we do not know.

Preeclampsia remains a major health problem for mothers and infants. Studying the entire pathophysiology of preeclampsia rather than "pregnancy-induced hypertension" has greatly expanded our knowledge of the disorder. Current thinking approaches preeclampsia as a 2 stage disorder: reduced placental perfusion usually secondary to abnormal implantation and a consequent maternal disorder characterized by endothelial dysfunction and subsequent pathophysiological changes. We know much about the 2 stages and less about their linkage. It is evident that reduced placental perfusion is not sufficient to account for the pathophysiology. Reduced perfusion and abnormal implantation occur in other conditions (intrauterine growth restriction and preterm labor) without the maternal syndrome. This leads to the hypothesis that reduced placental perfusion must interact with maternal constitutional factors to generate the systemic pathophysiology of preeclampsia. The similarities of these risk factors and metabolic alterations between preeclampsia and atherosclerosis suggest a common pathophysiology. Oxidative stress is postulated as the genesis of endothelial dysfunction in atherosclerosis. The author proposes that oxidative stress secondary to reduced placental perfusion leads to endothelial dysfunction, linking the 2 stages of the syndrome.

Animals↗

PubMed2000

Proteasomal turnover of p21Cip1 does not require p21Cip1 ubiquitination.

The Cdk inhibitor p21Cip1 is an unstable protein. Pharmacologic inhibition of the proteasome increases the half-life of p21 from less than 30 min to more than 2 hr and results in the accumulation of p21-ubiquitin conjugates. To determine whether ubiquitination was required for proteasomal degradation of p21, we constructed mutant versions of p21 that were not ubiquitinated in vivo. Remarkably, these mutants remained unstable and increased in abundance upon proteasome inhibition, indicating that direct ubiquitination of p21 is not necessary for its turnover by the proteasome. The frequently observed correlation between protein ubiquitination and proteasomal degradation is insufficient to conclude that ubiquitination is a prerequisite for degradation.

3T3 Cells↗

PubMed2000

Free leptin is increased in normal pregnancy and further increased in preeclampsia.

We measured bound and free leptin levels in preeclamptic and matched normal pregnant and never-pregnant women to determine whether the free component of leptin is increased during pregnancy and further increased in preeclampsia. Two milliliters of serum was obtained from 18 normal and 18 preeclamptic patients matched by pre-pregnancy body mass index (BMI), and from 18 never-pregnant women matched by BMI with the pregnant groups. The sample was subjected to gel filtration using Sephadex G-100. Radioimmunoassay (RIA) was performed on all fractions, and the proportions of bound and free leptin were determined by analyzing the areas under the curve of the chromatographic profile. The total maternal serum leptin concentration was significantly higher in normal pregnancy compared with the nonpregnant state and was further increased in preeclampsia (33.8 +/- 4.1 v 15.2 +/- 1.8 ng/mL, P = .002, and 48.1 +/- 5.6 ng/mL, P = .02, respectively). Free leptin was increased in normal pregnant compared with never-pregnant women (25.9 +/- 4.1 v 11.0 +/- 2.0 ng/mL, respectively, P = .01), while the increase of total leptin in preeclampsia was exclusively in the free fraction that was significantly higher versus the normal pregnant group (41.8 +/- 5.6 v25.9 +/- 4.1 ng/mL, respectively, P = .01). The bound leptin fraction, by contrast, was significantly increased in the normal pregnant group compared with the preeclamptic group and the never-pregnant group (7.9 +/- 0.56 v 6.2 +/- 0.36 and 4.1 +/- 0.36 ng/mL, respectively, P = .009 and P = <.0001). In conclusion, the free leptin concentration increases in normal pregnancy and is further increased in preeclampsia. This supports the hypothesis that biologically active leptin is elevated in normal pregnancy and is increased more in women with preeclampsia.

Adult↗

PubMed2000

Preterm prediction study: is socioeconomic status a risk factor for bacterial vaginosis in Black or in White women?

Bacterial vaginosis (BV), an important risk factor for preterm birth, is a more common infection in Black compared with White pregnant women. Because Black women in the United States are more likely to have lower measures of socioeconomic status (SES), this study examined the hypothesis that BV is associated with low SES. The project evaluated data from the Preterm Prediction Study of 2,929 women prospectively followed during their pregnancies. The women, who were screened for BV at 24 and 28 weeks of gestation, underwent a structured interview to evaluate demographic factors, SES, home and work environment, drug or alcohol use, and prior medical history. Black women in the study had many measures of lower SES compared with the White women, and reported less use of tobacco, alcohol and drugs. In neither the Black nor White women was an association found between BV and measures of SES (with the sole exception of "absence of a home telephone"). Most measures of SES do not explain the difference in rates of BV in Black and in White pregnant women.

Black or African American↗

PubMed2000

The preterm prediction study: granulocyte colony-stimulating factor and spontaneous preterm birth. National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network.

OBJECTIVE: Granulocyte colony-stimulating factor is elevated in the amniotic fluid and plasma of women with chorioamnionitis and active preterm labor. We investigated the relationship between plasma granulocyte colony-stimulating factor and subsequent spontaneous preterm birth in pregnant women without symptoms. STUDY DESIGN: We performed a nested case-control study involving 194 women who had a singleton spontaneous preterm birth and 194 matched term control subjects from the patient pool (n = 2929) enrolled in the Preterm Prediction Study. Plasma collected at 24 and 28 weeks' gestation was analyzed for granulocyte colony-stimulating factor, and the results were compared with subsequent spontaneous preterm birth. RESULTS: Compared with term control subjects, women who were delivered of their infants spontaneously at <28 weeks' gestation had increased mean granulocyte colony-stimulating factor values at 24 weeks' gestation (84.7 +/- 38.4 vs 67.7 +/- 28.6 pg/mL; P =.049), and women who were delivered of their infants at <32 weeks' gestation had increased mean plasma granulocyte colony-stimulating factor values at 28 weeks' gestation (80.4 +/- 24.1 vs 55.9 +/- 16.5 pg/mL; P =. 001). At 24 weeks' gestation a granulocyte colony-stimulating factor value >75th percentile in control subjects (approximately 80 pg/mL) was found in 48.9% (23/47) of all women delivered of their infants at <32 weeks' gestation versus 14.9% (7/47) of the term control subjects (adjusted odds ratio, 6.2; 95% confidence interval, 1.8-20. 8). At 28 weeks' gestation a granulocyte colony-stimulating factor value >75th percentile was found in 36.8% (7/19) of women delivered of their infants at <32 weeks' gestation versus 5.3% (1/19) of term control subjects (adjusted odds ratio, 25.7; 95% confidence interval, 1.5-470.4). When measured at 24 or 28 weeks' gestation, granulocyte colony-stimulating factor did not predict spontaneous preterm birth at 32 to 34 weeks' gestation or at 35 to 36 weeks' gestation. CONCLUSION: In pregnant women without symptoms at 24 and 28 weeks' gestation, elevated plasma granulocyte colony-stimulating factor levels are associated with subsequent early (<32 weeks' gestation) spontaneous preterm birth, especially within the next 4 weeks, but not with late spontaneous preterm birth. These data provide further evidence that early spontaneous preterm birth is associated with an inflammatory process that is identifiable by the presence of a cytokine in maternal plasma several weeks before the early spontaneous preterm birth; however, later spontaneous preterm birth is not associated with this process.

Adolescent↗

PubMed2000

The preterm prediction study: cervical lactoferrin concentration, other markers of lower genital tract infection, and preterm birth. National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network.

OBJECTIVE: This study was undertaken to determine the relationship among cervical lactoferrin concentration, other cervical markers potentially related to infection, and spontaneous preterm birth. STUDY DESIGN: Cervical lactoferrin concentrations obtained at 22 to 24 weeks' gestation among 121 women who had a spontaneous preterm birth <35 weeks' gestation were compared with cervical lactoferrin concentrations among 121 women matched for race, parity, and center who were delivered at >/=37 weeks' gestation. Results were compared against levels of cervical interleukin 6, fetal fibronectin, and sialidase, against cervical length according to ultrasonography, and according to the bacterial vaginosis Gram stain score. RESULTS: Cervical lactoferrin concentrations ranged from not measurable (19% of the concentrations were below the threshold for this assay) to a titer of >/=1:64. There was no significant difference in the overall distributions of lactoferrin concentrations between the case patients and control subjects (P =.18). Only when the highest titers of lactoferrin were considered were there more women in the spontaneous preterm birth group (6/121 vs 0/121; P =.03). According to Spearman correlation analyses the cervical lactoferrin concentrations were strongly related to interleukin 6 concentration (r =.51; P =.0001), sialidase activity (r =.38; P =.0001), and bacterial vaginosis (r =.38; P =.0001), were weakly related to fetal fibronectin (r =. 16; P =.01), and were not related to cervical length. With the 90th percentile (a dilution of 1:32) used as a cutoff to establish a dichotomous variable, lactoferrin concentration had the following odds ratios and 95% confidence intervals for associations with other potential markers of infection: bacterial vaginosis odds ratio, 4.8 (95% confidence interval, 2.2-10.3); interleukin 6 concentration odds ratio, 2.8 (95% confidence interval, 1.2-6.5); sialidase activity odds ratio, 5. 5 (95% confidence interval, 2.2-13.7); fetal fibronectin concentration odds ratio, 0.6 (95% confidence interval, 0.2-2.0); chlamydiosis odds ratio, 2.3 (95% confidence interval, 0.8-6.9); and short cervix odds ratio, 0.5 (95% confidence interval, 0.2-1.4). CONCLUSIONS: Lactoferrin found in the cervix correlated well with other markers of lower genital tract infection. High lactoferrin levels were associated with spontaneous preterm birth but had a very low predictive sensitivity.

Bacterial Infections↗

PubMed2000

The Preterm Prediction Study: sequential cervical length and fetal fibronectin testing for the prediction of spontaneous preterm birth. National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network.

OBJECTIVES: This study was undertaken to further elucidate the pathogenesis of preterm birth by means of traditional risk factors and new markers for preterm birth derived from the Preterm Prediction Study. STUDY DESIGN: A total of 3076 women (2929 with singleton gestations and 147 with twin pregnancies) were categorized according to the presence of risk factors including black race, low body mass index, the presence of bacterial vaginosis, and previous preterm birth. At 24 and 28 weeks' gestation cervical length was measured and categorized as short (</=25 mm) or normal (>25 mm). Vaginal and cervical fetal fibronectin concentrations were measured at 24, 26, 28, and 30 weeks' gestation and results were categorized as positive (>/=50 ng/mL) or negative (<50 ng/mL). RESULTS: At 24 to 26 weeks' gestation women with each of the risk factors were more likely to have positive fibronectin test results or to have a short cervix. Among women with negative fetal fibronectin results at 24 to 26 weeks' gestation those with a short cervix were more likely to have positive fetal fibronectin results at 28 to 30 weeks' gestation, and among those with normal cervical length those women who had positive fetal fibronectin results were more likely to have a short cervix at later evaluation. Most women who had positive fetal fibronectin results at 24 to 26 weeks' gestation had negative results at 28 to 30 weeks' gestation, whereas most but not all women who had a short cervix at 24 to 26 weeks' gestation still had a short cervix at 28 to 30 weeks' gestation. In each period women with both a positive fetal fibronectin result and a short cervix were at substantially increased risk of spontaneous preterm birth; women with either marker alone had intermediate and approximately equal risks of spontaneous preterm birth, and women without either marker had a low risk of spontaneous preterm birth. CONCLUSION: Regardless of other risk factors, a short cervix predicts a subsequent positive fetal fibronectin result, and a positive fetal fibronectin result predicts subsequent cervical shortening. These data do not support a single sequence of events leading to spontaneous preterm birth.

Biomarkers↗

PubMed2000

Vaginal fetal fibronectin measurements from 8 to 22 weeks' gestation and subsequent spontaneous preterm birth.

OBJECTIVE: We sought to determine the range of fetal fibronectin values in the vagina from 8 to 22 weeks' gestation, the factors associated with both low and high values, and whether high values are associated with gestational age at birth. STUDY DESIGN: Vaginal fetal fibronectin was quantitatively determined in a prospective cohort study of 13,360 women being evaluated for participation in the National Institute of Child Health and Human Development Maternal-Fetal Medicine Unit treatment trials for bacterial vaginosis and Trichomonas vaginalis. Fetal fibronectin values were correlated with gestational age at screening, race, the presence of bacterial vaginosis and Trichomonas vaginalis, and gestational age at delivery. RESULTS: Vaginal fetal fibronectin values at each gestational age ranged from unmeasurable to >1000 ng/mL, with median values always being <10 ng/mL. Fetal fibronectin values declined progressively with increasing gestational age at sampling. Bacterial vaginosis and black race were associated with higher values, whereas nulliparity was associated with lower values. High values after 13 weeks' gestation were associated with a 2- to 3-fold increased risk of subsequent spontaneous preterm birth overall and a 4-fold increased risk of very early preterm birth. CONCLUSION: Elevated vaginal fetal fibronectin levels from 13 to 22 weeks' gestation are associated with a significantly increased risk of spontaneous preterm birth.

Black People↗

PubMed2000

The impact of digital cervical examination on expectantly managed preterm rupture of membranes.

OBJECTIVE: The purpose of this study was to examine the effects of digital cervical examination on maternal and neonatal outcomes among women with preterm rupture of membranes. STUDY DESIGN: This analysis includes data from a previously reported trial of antibiotic treatment during expectant management of rupture of membranes at 24 to 32 weeks' gestation in singleton and twin gestations. Patients from both the randomized trial (n = 299 in the antibiotic group and n = 312 in the placebo group) and the observational component (n = 183) are included in this analysis. The groups were divided into those with one (n = 161) or two digital cervical examinations (n = 27) and those with no digital cervical examinations (n = 606). RESULTS: The gestational ages at enrollment were similar in the two groups (29 +/- 2 weeks' gestation for one or two examinations vs 29 +/- 2 weeks' gestation for no examinations; P =.85). There were no differences in chorioamnionitis (27% vs 29%; P =.69), endometritis (13% vs 11%; P =.5), or wound infection (0.5% vs 1%; P >.999) between the group with one or two examinations and the no-examination group. Infant outcomes were also similar in the two groups, including early sepsis (6% vs 5%; P =.68), respiratory distress syndrome (51% vs 45%; P =.18), intraventricular hemorrhage (7% vs 7%; P =.67), necrotizing enterocolitis (5% vs 3%; P =.19), and perinatal death (7% vs 5%; P =.45). A composite outcome made up of these neonatal outcomes was not different (56% vs 48%; P =.10) between the group with one or two examinations and the no-examination group. The time from rupture to delivery was shorter in the digital examination group (median value, 3 vs 5 days; P <. 009). Multivariable analysis to adjust for antibiotic study group, group B streptococcal culture status, race, and maternal transfer did not modify these results. CONCLUSION: Performance of one or two digital cervical examinations during the course of expectant management of rupture of membranes between 24 and 32 weeks' gestation was associated with shorter latency but did not appear to worsen either maternal or neonatal outcome.

Adult↗

PubMed2000

The Preterm Prediction Study: prediction of preterm premature rupture of membranes through clinical findings and ancillary testing. The National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network.

OBJECTIVE: Our objective was to determine the relative importance of demographic characteristics, clinical risk factors, and ancillary screening tests in the prediction of preterm birth as a result of premature rupture of membranes. STUDY DESIGN: A total of 2929 women were evaluated in 10 centers at 23 to 24 weeks' gestation. Demographic and clinical characteristics were ascertained. Cervicovaginal fetal fibronectin and bacterial vaginosis were evaluated. Cervical length was measured by vaginal ultrasonography. Patients were evaluated for spontaneous preterm birth caused by preterm premature rupture of membranes at <37 and <35 weeks' gestation. Multivariate analyses were performed separately for nulliparous women and multiparous women. RESULTS: Premature rupture of membranes at <37 weeks' gestation complicated 4.5% of pregnancies, accounting for 32.6% of preterm births. Univariate analysis revealed low body mass index, pulmonary disease, contractions within 2 weeks, short cervix (</=25 mm), positive results of fetal fibronectin screening, bacterial vaginosis, and a previous preterm birth caused by premature rupture of membranes (in multiparous women) to be significantly associated with preterm birth caused by premature rupture of membranes in the current gestation. Short cervix, previous preterm birth caused by premature rupture of membranes in multiparous women, and presence of fetal fibronectin were the strongest predictors for both preterm birth caused by premature rupture of membranes at <37 and <35 weeks' gestation. Women with positive fetal fibronectin screening results and a short cervix had greater risks both of preterm birth caused by premature rupture of membranes at <37 weeks' gestation (relative risk, 4.9) and of preterm birth caused by premature rupture of membranes at <35 weeks' gestation (relative risk, 13.5) than did those without these risk factors. Multiparous women with all three risk factors had a 31. 3-fold increased risk of preterm birth caused by premature rupture of membranes at <35 weeks' gestation. CONCLUSION: The combination of short cervical length, previous preterm birth caused by preterm premature rupture of membranes, and positive fetal fibronectin screening results was highly associated with preterm delivery caused by preterm premature rupture of membranes in the current gestation.

Cervix Uteri↗

PubMed2000

The preterm prediction study: quantitative fetal fibronectin values and the prediction of spontaneous preterm birth. The National Institute of Child Health and Human Development Maternal-Fetal Medicine Units Network.

OBJECTIVE: A cervicovaginal fetal fibronectin value of >/=50 ng/mL has been used to define women at risk of having a preterm birth. We evaluated the relationship between quantitative fetal fibronectin values and spontaneous preterm birth. STUDY DESIGN: Cervical and vaginal specimens for fetal fibronectin were obtained at 24, 26, 28, and 30 weeks' gestation from 2926 women. Quantitative fetal fibronectin values were calculated by using absorbances determined by enzyme-linked immunosorbent assay. The highest fetal fibronectin value (cervical or vaginal) for each woman at each visit was evaluated in relation to spontaneous preterm birth at <35 weeks' gestation. Receiver operating characteristic curves were constructed to determine the optimal cutoff point for fetal fibronectin values to predict spontaneous preterm birth at <35 weeks' gestation and within 4 weeks of testing. RESULTS: The risk of spontaneous preterm birth increased as a function of increasing fetal fibronectin values from approximately 20 to 300 ng/mL. Fetal fibronectin values > or =300 ng/mL were not associated with a further increase in spontaneous preterm birth. Examination of the receiver operating characteristic curve indicates that the optimal cutoff point for a positive fetal fibronectin test result at 24 to 30 weeks' gestation to predict spontaneous preterm birth at <35 weeks is between 45 and 60 ng/mL. CONCLUSION: Increasing levels of cervicovaginal fetal fibronectin up to 300 ng/mL are associated with an increasing risk of spontaneous preterm birth. Nevertheless, at 24 to 30 weeks, the value currently used, 50 ng of fetal fibronectin per milliliter, appears to be a reasonable cutoff point for predicting spontaneous preterm birth at <35 weeks' gestation.

Cervix Uteri↗

PubMed2000

Risk factors for community-acquired pneumonia among persons infected with human immunodeficiency virus.

Two hundred eleven adults with human immunodeficiency virus (HIV) infection hospitalized for community-acquired pneumonia, including Pneumocystis carinii pneumonia (PCP; patients), and 192 matched HIV-infected hospitalized patients without pneumonia (controls) were interviewed to determine risk factors for pneumonia. Multivariate logistic regression showed that patients were less likely than controls to have used trimethoprim-sulfamethoxazole (TMP-SMZ) prophylaxis (odds ratio [OR], 0.22; 95% confidence interval [CI], 0.12-0.41) and more likely to have been hospitalized previously with pneumonia (OR, 6.25; CI, 3.40-11.5). Patients were also more likely than controls to have gardened (OR, 2.24; CI, 1.00-5.02) and to have camped or hiked (OR, 4.95; CI, 1.31-18.7), but stratified analysis by etiologic agent showed this association only for PCP. These findings reconfirm the efficacy of TMP-SMZ in preventing community-acquired pneumonia. In addition, hospitalization for pneumonia might represent a missed opportunity to encourage HIV-infected patients to enter into regular medical care and to adhere to prescribed antiretroviral and prophylaxis medications.

Adult↗

PubMed2000

An unusual case of unilateral lactation and a vanishing limb.

Explore the source record for details and available documents.

Adult↗

PubMed2000

Dyslipoproteinaemia in postmenopausal women with a history of eclampsia.

OBJECTIVE: To test the hypothesis that postmenopausal women with a history of eclampsia manifest a more high risk lipid profile than postmenopausal women with a history of normal pregnancy. SETTING: The Department of Obstetrics and Gynaecology, National University Hospital, Reykjavik, Iceland, and the Magee-Womens Research Institute, Pittsburgh, Pennsylvania, USA. PARTICIPANTS: Thirty Icelandic women with a history of eclampsia, aged between 50 and 67 years at the time of re-examination (cases) were individually matched for current age, and for age and parity at index pregnancy, to 30 unrelated Icelandic women with a history of normal pregnancy (controls). METHODS: The participating women completed a health and family history questionnaire and underwent a physical examination. Fasting plasma low density lipoprotein diameter, serum lipids, insulin, and glucose were measured. RESULTS: Mean low density lipoprotein size was significantly smaller and apolipoprotein B concentration was higher in women with prior eclampsia. The percentage of cases receiving blood pressure medication (33%) was significantly greater than controls (6.7%). Thirteen cases had had hypertensive complications in at least one other pregnancy (recurrent subgroup); postmenopausally, these women displayed significantly increased diastolic blood pressures, smaller-sized low density lipoprotein, increased apolipoprotein B, decreased high density lipoprotein2 (HDL2) cholesterol, and increased total cholesterol: HDL cholesterol ratio compared with their controls. Fourteen cases were normotensive in all other pregnancies (nonrecurrent); these showed no differences from their controls. CONCLUSIONS: Dyslipoproteinaemia is more prevalent among postmenopausal women with prior eclampsia, especially with recurrent hypertension in pregnancy, than in postmenopausal women with prior normal pregnancies.

Aged↗

PubMed2000

Mild gait abnormality and leg discomfort in a child secondary to extradural ganglioneuroma.

Ganglioneuromas are benign and slow-growing tumors that most commonly originate from the sympathetic trunk. Ganglioneuromas often decrease in size and rarely require reoperation. Changes in gait or the onset of limb pain without a discernible local cause are indications for investigation of patients for possible intraspinal pathology. We report the case of a 5-year-old boy who presented with seemingly static symptoms, while the slow-growing tumor had enveloped nerve roots and caused bone destruction of the vertebrae.

Biopsy↗

PubMed2000

Developing new competencies in healthcare practitioners in the field.

The Life-long Learning concept is one which is appropriate for those who have (or create) the opportunities to develop their competencies once they are in an operational role. All models of healthcare delivery and management are VOLATILE and therefore the way of addressing new competencies cannot be prescriptive or stand still for too long, but the concepts do endure. Learning, in order to 'keep up to date' is very necessary--technology, new clinical practices and interventions, new drug interactions, increased patient demands ... all add to the need to move with the times. This paper addresses some of the issues surrounding this challenge in the domain of informatics in support of healthcare delivery and management.

Competency-Based Education↗

PubMed1999

Cullin-3 targets cyclin E for ubiquitination and controls S phase in mammalian cells.

Cyclin E is an unstable protein that is degraded in a ubiquitin- and proteasome- dependent pathway. Two factors stimulate cyclin E ubiquitination in vivo: when it is free of its CDK partner, and when it is phosphorylated on threonine 380. We pursued the first of these pathways by using a two-hybrid screen to identify proteins that could bind only to free cyclin E. This resulted in the isolation of human Cul-3, a member of the cullin family of E3 ubiquitin-protein ligases. We found that Cul-3 was bound to cyclin E but not to cyclin E-Cdk2 complexes in mammalian cells, and that overexpression of Cul-3 increased ubiquitination of cyclin E but not other cyclins. Conversely, deletion of the Cul-3 gene in mice caused increased accumulation of cyclin E protein, and had cell-type-specific effects on S-phase regulation. In the extraembryonic ectoderm, in which cells undergo a standard mitotic cycle, there was a greatly increased number of cells in S phase. In the trophectoderm, in which cells go through endocycles, there was a block to entry into S phase. The SCF pathway, which targets cyclins for ubiquitination on the basis of their phosphorylation state, and the Cul-3 pathway, which selects cyclin E for ubiquitination on the basis of its assembly into CDK complexes, may be complementary ways to control cyclin abundance.

3T3 Cells↗