Assessing and Treating Cardiovascular Risk: Traditional Moderation is the Way to Go.

No abstract available.

Interorganellar signal transduction: the arrest of secretion response.

Signals from the secretory pathway caused by "traffic jams" are transduced by the WSC1, WSC2/PKC1 signaling pathway and result in nuclear reorganization and ribosomal gene repression.

Distinct retrieval and retention mechanisms are required for the quality control of endoplasmic reticulum protein folding.

Proteins destined for the secretory pathway must first fold and assemble in the lumen of endoplasmic reticulum (ER). The pathway maintains a quality control mechanism to assure that aberrantly processed proteins are not delivered to their sites of function. As part of this mechanism, misfolded proteins are returned to the cytosol via the ER protein translocation pore where they are ubiquitinated and degraded by the 26S proteasome. Previously, little was known regarding the recognition and targeting of proteins before degradation. By tracking the fate of several mutant proteins subject to quality control, we demonstrate the existence of two distinct sorting mechanisms. In the ER, substrates are either sorted for retention in the ER or are transported to the Golgi apparatus via COPII-coated vesicles. Proteins transported to the Golgi are retrieved to the ER via the retrograde transport system. Ultimately, both retained and retrieved proteins converge at a common machinery at the ER for degradation. Furthermore, we report the identification of a gene playing a novel role specific to the retrieval pathway. The gene, BST1, is required for the transport of misfolded proteins to the Golgi, although dispensable for the transport of many normal cargo proteins.

Prospective study of 1 surgeon's experience with 115 cases using the Unfolder lens injection system.

PURPOSE: To describe the experience of 1 surgeon's first 115 cases using the Allergan Unfolder intraocular lens (IOL) injection system. SETTING: Free-standing day-surgery center, Chatswood, Australia. METHODS: After standard phacoemulsification, an SI-40NB IOL (Allergan) was implanted using the Unfolder. Data on complications and the success of implantation were collected prospectively. RESULTS: Ninety-four percent of patients achieved a 1 month visual acuity of 6/6 or better. Of the 13 cases with a potential complication related to the Unfolder, all achieved a visual acuity of 6/6 or better. No complication affected the eye adversely or reduced visual acuity significantly. CONCLUSION: Implantation of SI-40NB IOLs with the Unfolder led to no serious complications and gave satisfactory visual results.

The unfolded protein response: no longer just a special teams player.

The endoplasmic reticulum stress pathway known as the unfolded protein response is currently the best understood model of interorganellar signal transduction. Bridging a physical separation, the pathway provides a direct line of communication between the endoplasmic reticulum lumen and the nucleus. With the unfolded protein response, the cell has the means to monitor and respond to the changing needs of the endoplasmic reticulum. Beginning with the discovery of its remarkable signaling mechanism in yeast, the unfolded protein response has not ceased to reveal more of its many secrets. By applying powerful biochemical, genetic, genomic, and cytological approaches, the recent efforts of many groups have buried the long-held notion that the unfolded protein response is simply a regulatory platform for endoplasmic reticulum chaperones. We now know that the unfolded protein response regulates many genes that affect diverse aspects of cellular physiology. In addition, studies in mammals have revealed novel unfolded protein response signaling factors that may contribute to the specialized needs of multicellular organisms. This article focuses on these and other recent developments in the field.

The unfolded protein response regulates multiple aspects of secretory and membrane protein biogenesis and endoplasmic reticulum quality control.

The unfolded protein response (UPR) is an intracellular signaling pathway that relays signals from the lumen of the ER to activate target genes in the nucleus. We devised a genetic screen in the yeast Saccharomyces cerevisiae to isolate mutants that are dependent on activation of the pathway for viability. Using this strategy, we isolated mutants affecting various aspects of ER function, including protein translocation, folding, glycosylation, glycosylphosphatidylinositol modification, and ER-associated protein degradation (ERAD). Extending results gleaned from the genetic studies, we demonstrate that the UPR regulates trafficking of proteins at the translocon to balance the needs of biosynthesis and ERAD. The approach also revealed connections of the UPR to other regulatory pathways. In particular, we identified SON1/RPN4, a recently described transcriptional regulator for genes encoding subunits of the proteasome. Our genetic strategy, therefore, offers a powerful means to provide insight into the physiology of the UPR and to identify novel genes with roles in many aspects of secretory and membrane protein biogenesis.

An evaluation of the implementation of case management in the community psychiatric nursing service.

This paper reports the preliminary results of a study on the implementation of case management in the community psychiatric nursing service (CPNS). The purpose of the study is to compare the outcome of case management service with the conventional practice of CPNS in the care of chronic schizophrenic clients. The impact of case management service on clients' clinical status, functional level and satisfaction was measured. A matched, pre-post, case-control group design was used to compare case management and conventional care groups. A specific case management model based on the practice at Carondelet Saint Mary's Hospital and Health Centre in Tucson has been developed to care for the schizophrenic clients in the community. Subjects in the experimental group were cared for by CPNs using this case management model. Subjects in the control group were cared for by CPNs based on current practices in the CPNS. Pre- and post-measures were taken upon recruitment and at 5 months later. Findings showed that the experimental group had better outcome in terms of their mental status and functional level when compared with the control group. They were generally more satisfied with the service. When comparing case managers' role with that of conventional CPNs, case managers performed more in areas such as assessment, liaison, case discussion and life skills training. This study demonstrates that this particular model of case management can be effective in caring for clients with long-term mental health problems in Hong Kong.

Case management in the Community Psychiatric Nursing Service in Hong Kong: describing the process.

PURPOSE: To explore the roles, functions, and work practices of case managers and their perceptions on the implementation of case management in the Community Psychiatric Nursing Services. METHODS: A qualitative descriptive approach was used. Data were collected from daily reports, field observations, reflective diaries, and small group interviews. FINDINGS: The study showed the work practices of case managers were more intensive compared to the conventional community psychiatric nursing practice, and there was more involvement of clients and their family in the treatment process. Case managers identified liaison, coordination, supportive and family counseling, and teaching practical living skills as their important roles. CONCLUSIONS: This study provides valuable information in understanding the process of case management and contributed to the development of case management as a care delivery model in Hong Kong.

The signal for clathrin-mediated endocytosis of the paramyxovirus SV5 HN protein resides at the transmembrane domain-ectodomain boundary region.

The hemagglutinin-neuraminidase (HN) glycoprotein of the paramyxovirus SV5 is internalized from the cell surface via clathrin-coated pits. However, the cytoplasmic domain of SV5 HN does not contain a previously characterized internalization motif. A cell-surface-expressed chimeric protein (APK), consisting of the cytoplasmic tail, transmembrane (TM) domain, and 12 residues of the ectodomain of HN joined to the cytoplasmic protein pyruvate kinase is internalized, indicating that the N-terminal region of HN contains an internalization signal. Although SV5 HN is internalized at a rate similar to that of influenza virus hemagglutinin (HA) mutant Y543, which contains a degenerate tyrosine-based signal in its cytoplasmic tail, the elimination of the majority of the HN cytoplasmic tail, or substitution of the HN TM domain with leucine residues, did not affect the rate of HN internalization. The HN protein of the closely related virus, Newcastle disease virus (NDV), is not internalized from the cell surface. Working under the usual convention that the TM domain consists of the hydrophobic residues bounded by two charged residues, analysis of internalization of mutant and chimeric NDV HN molecules indicates that the first seven SV5 HN ectodomain residues are critical for internalization of HN. A glutamic acid residue (E37) that abuts this presumptive HN TM domain/ectodomain boundary is important for SV5 HN internalization.

Intraoperative complications of 1000 phacoemulsification procedures: a prospective study.

PURPOSE: To determine the frequency and nature of intraoperative complications of endocapsular phacoemulsification cataract surgery. SETTING: Dalcross Private Hospital and the Departments of Ophthalmology, Concord Hospital and Prince of Wales Hospital, Sydney, Australia. METHODS: This prospective study included the first 1000 cases of planned endocapsular phacoemulsification cataract surgery performed by an experienced surgeon. A standardized proforma was completed at the time of surgery. Data recorded included pre-existing ocular abnormalities, duration of surgery, nuclear sclerosis grade, and intraoperative complications. RESULTS: Major complications comprised posterior capsule tears with vitreous loss (1.4%), isolated posterior capsule tears (0.7%), and zonulysis (0.1%). Minor complications included anterior capsule tears (3.8%), iris prolapse (0.6%), and ciliary body incision (0.2%). There was a significant trend toward complications as the nuclear sclerosis grade increased. The incidence of major complications was 9.3% in the first 150 cases and 0.9% in the last 850. This represents a relative risk of 9.9 (95% confidence interval 4.2 to 23.0) of a major complication occurring in the first 150 compared with the later 850 cases. CONCLUSION: The frequency of major and minor complications fell sharply after the first 150 operations and was maintained. This study may provide a guide for beginning phacoemulsification surgeons and a basis for experienced surgeons to compare their performance outcomes.

Effects of sorbitol dehydrogenase deficiency on nerve conduction in experimental diabetic mice.

In this report, we made use of sorbitol dehydrogenase (SDH)-deficient mutant mice (C57BL/LiA) to test whether there is a close correlation between the level of polyol accumulation and the degree of reduction in motor nerve conduction velocity (MNCV) associated with diabetes. The C57BL/LiA mouse has SDH deficiency due to a G-to-A mutation at the +1 position of intron 8, thus producing only aberrant SDH transcripts. These C57BL/LiA mice should have higher levels of polyol accumulation in the peripheral nerve because of the inability to further metabolize sorbitol to fructose. Here, we confirm by Western blot analysis and high-performance liquid chromatography that these mice lack SDH in the sciatic nerve and other various tissues, whereas normal mice possess SDH. These C57BL/LiA mice do not display any obvious phenotype that includes peripheral neuropathy in the normal laboratory environment and breed normally as described previously, although the tissues that normally contain SDH accumulate more sorbitol. This finding suggested that C57BL/LiA mouse strain is a valid model for studying the role in diabetic neuropathy of the polyol pathway, which consists of two enzymes-aldose reductase for converting glucose to sorbitol and SDH for converting sorbitol to fructose. Sorbitol levels in the sciatic nerve of diabetic C57BL/10N, nondiabetic, and diabetic C57BL/LiA mice were increased 4.3-, 16.6-, and 38.1-fold, respectively, above that of nondiabetic C57BL/10N. The fructose level in the sciatic nerve was increased 2.4-fold in diabetic C57BL/10N mice compared with that of nondiabetic and diabetic C57BL/LiA mice. Diabetic SDH-deficient mice showed an MNCV reduction similar in magnitude to that of diabetic C57BL/10N mice, despite greater nerve sorbitol accumulation and the lack of fructose in the former. The present data suggest that the levels of sorbitol and fructose in the sciatic nerve of mice do not correlate with the severity of MNCV deficit associated with diabetes.

Signal sequences specify the targeting route to the endoplasmic reticulum membrane.

In the yeast Saccharomyces cerevisiae, only a subset of preproteins that are translocated across the ER membrane require the function of the signal recognition particle (SRP), suggesting that an alternative, SRP-independent pathway must exist (Hann, B.C., and P. Walter. 1991. Cell. 67:131-144). We have established that the two targeting pathways function in parallel. Mutant alleles of SEC62 and SEC63 were isolated that specifically impaired the translocation of SRP-independent preproteins in vivo and in vitro, whereas SRP-dependent preproteins were unaffected. Based on this analysis, preproteins fall into three distinct classes: SRP dependent, SRP independent, and those that can use both pathways. Pathway specificity is conferred by the hydrophobic core of signal sequences. Our studies show a previously unrecognized diversity in ER-directed signal sequences, that carry structural information that serves to identify the route taken.

ER membrane protein complex required for nuclear fusion.

Diploid cells of the yeast Saccharomyces cerevisiae form after the mating of two haploid cells of the opposite mating type. After fusion of the two plasma membranes of the mating cells, a dinucleated cell forms initially in which the two haploid nuclei then rapidly fuse to form a single diploid nucleus. This latter event, called karyogamy, can be divided into two distinct steps: the microtubule-based movement that causes the two nuclei to become closely juxtaposed and the fusion of the nuclear membranes. For the membrane fusion step, one required component, the ER luminal protein Kar2p (BiP), has been identified. For topological reasons, however, it has been unclear how Kar2p could function in this role. Kar2p is localized to the luminal (i.e., noncytoplasmic) face of the ER membrane, yet nuclear fusion must initiate from the cytosolic side of the outer nuclear membrane or the ER membrane with which it is contiguous. There is both genetic and biochemical evidence that Kar2p interacts with Sec63p, an ER membrane protein containing both luminal and cytosolic domains that is involved in protein translocation across the membrane. We have isolated novel sec63 mutant alleles that display severe karyogamy defects. Disruption of the genes encoding other Sec63p-associated proteins (Sec71p and Sec72p) also results in karyogamy defects. A suppressor mutant (sos1-1) partially corrects the translocation defect but does not alleviate the karyogamy defect. sec61 and sec62 mutant alleles that cause similar or more severe protein translocation defects show no karyogamy defects. Taken together, these results suggest a direct role for Sec63p, Sec71p, and Sec72p in nuclear membrane fusion and argue against the alternative interpretation that the karyogamy defects result as an indirect consequence of the impaired membrane translocation of another component(s) required for the process. We propose that an ER/nuclear membrane protein complex composed of Sec63p, Sec71p, and Sec72p plays a central role in mediating nuclear membrane fusion and requires ER luminally associated Kar2p for its function.

Protein translocation across the endoplasmic reticulum.

In the past year, dramatic progress has been made in our understanding of protein biogenesis at the initial steps of the eukaryotic secretory pathway. New insights have refined our view of protein targeting to the endoplasmic reticulum membrane and provided the best glimpse so far of the subsequent translocation step. The interactions of three GTP-binding proteins have been found to result in a novel cycle of GTP binding and hydrolysis to regulate protein targeting. Experiments with fluorescent probes have revealed that the nascent chain enters an aqueous environment within the membrane sealed off from the cytosol. In vitro reconstitution experiments have shown surprising simplicity in the number of polypeptides required to facilitate translocation across a synthetic membrane and to promote the integration of membrane proteins. Furthermore, new genetic and functional similarities between divergent organisms have been discovered, providing convincing evidence of the evolutionary conservation of strategies used in the targeting and translocation of polypeptides.

Analysis in vivo of GRP78-BiP/substrate interactions and their role in induction of the GRP78-BiP gene.

The endoplasmic reticulum (ER)-localized chaperone protein, GRP78-BiP, is involved in the folding and oligomerization of secreted and membrane proteins, including the simian virus 5 hemagglutinin-neuraminidase (HN) glycoprotein. To understand this interaction better, we have constructed a series of HN mutants in which specific portions of the extracytoplasmic domain have been deleted. Analysis of these mutant polypeptides expressed in CV-1 cells have indicated that GRP78-BiP binds to selective sequences in HN and that there exists more than a single site of interaction. Mutant polypeptides have been characterized that are competent and incompetent for association with GRP78-BiP. These mutants have been used to show that the induction of GRP78-BiP synthesis due to the presence of nonnative protein molecules in the ER is dependent on GRP78-BiP complex formation with its substrates. These studies have implications for the function of the GRP78-BiP protein and the mechanism by which the gene is regulated.

Different roles of individual N-linked oligosaccharide chains in folding, assembly, and transport of the simian virus 5 hemagglutinin-neuraminidase.

The role of N-linked glycosylation in protein maturation and transport has been studied by using the simian virus 5 hemagglutinin-neuraminidase (HN) protein, a model class II integral membrane glycoprotein. The sites of N-linked glycosylation on HN were identified by eliminating each of the potential sites for N-linked glycosylation by oligonucleotide-directed mutagenesis on a cDNA clone. Expression of the mutant HN proteins in eucaryotic cells indicated that four sites are used in the HN glycoprotein for the addition of N-linked oligosaccharide chains. These functional glycosylation sites were systematically eliminated in various combinations from HN to form a panel of mutants in which the roles of individual carbohydrate chains and groups of carbohydrate chains could be analyzed. Alterations in the normal glycosylation pattern resulted in the impairment of HN protein folding and assembly which, in turn, affected the intracellular transport of HN. The severity of the consequences on HN maturation depended on both the number of deleted carbohydrate sites and their position in the HN molecule. Analysis of the reactivity pattern of HN conformation-specific monoclonal antibodies with the mutant HN proteins indicated that one specific carbohydrate chain plays a major role in promoting the correct folding of HN. Another carbohydrate chain, which is not essential for the initial folding of HN was found to play a role in preventing the aggregation of HN oligomers. The HN molecules which were misfolded, owing to their altered glycosylation pattern, were retained in the endoplasmic reticulum. Double-label immunofluorescence experiments indicate that misfolded HN and folded HN are segregated in the same cell. Misfolded HN forms disulfide-linked aggregates and is stably associated with the resident endoplasmic reticulum protein, GRP78-BiP, whereas wild-type HN forms a specific and transient complex with GRP78-BiP during its folding process.