Mutations in LMAN1 (also called ERGIC-53) result in combined deficiency of factor V and factor VIII (F5F8D), an autosomal recessive bleeding disorder characterized by coordinate reduction of both clotting proteins. LMAN1 is a mannose-binding type 1 transmembrane protein localized to the endoplasmic reticulum-Golgi intermediate compartment (ERGIC; refs. 2,3), suggesting that F5F8D could result from a defect in secretion of factor V and factor VIII (ref. 4). Correctly folded proteins destined for secretion are packaged in the ER into COPII-coated vesicles, which subsequently fuse to form the ERGIC. Secretion of certain abundant proteins suggests a default pathway requiring no export signals (bulk flow; refs. 6,7). An alternative mechanism involves selective packaging of secreted proteins with the help of specific cargo receptors. The latter model would be consistent with mutations in LMAN1 causing a selective block to export of factor V and factor VIII. But approximately 30% of individuals with F5F8D have normal levels of LMAN1, suggesting that mutations in another gene may also be associated with F5F8D. Here we show that inactivating mutations in MCFD2 cause F5F8D with a phenotype indistinguishable from that caused by mutations in LMAN1. MCFD2 is localized to the ERGIC through a direct, calcium-dependent interaction with LMAN1. These findings suggest that the MCFD2-LMAN1 complex forms a specific cargo receptor for the ER-to-Golgi transport of selected proteins.

Factor VIII (FVIII) functions as a cofactor within the intrinsic pathway of blood coagulation. Quantitative or qualitative deficiencies of FVIII result in the inherited bleeding disorder hemophilia A. Expression of FVIII (domain structure A1-A2-B-A3-C1-C2) in heterologous mammalian systems is 2 to 3 orders of magnitude less efficient compared with other proteins of similar size compromising recombinant FVIII production and gene therapy strategies. FVIII expression is limited by unstable mRNA, interaction with endoplasmic reticulum (ER) chaperones, and a requirement for facilitated ER to Golgi transport through interaction with the mannose-binding lectin LMAN1. Bioengineering strategies can overcome each of these limitations. B-domain-deleted (BDD)-FVIII yields higher mRNA levels, and targeted point mutations within the A1 domain reduce interaction with the ER chaperone immunoglobulin-binding protein. In order to increase ER to Golgi transport we engineered several asparagine-linked oligosaccharides within a short B-domain spacer within BDD-FVIII. A bioengineered FVIII incorporating all of these elements was secreted 15-to 25-fold more efficiently than full-length FVIII both in vitro and in vivo. FVIII bioengineered for improved secretion will significantly increase potential for success in gene therapy strategies for hemophilia A as well as improve recombinant FVIII production in cell culture manufacturing or transgenic animals. IntroductionFactor VIII (FVIII) is a large plasma glycoprotein that functions as an essential cofactor for the proteolytic activation of factor X by activated factor IX within the intrinsic pathway of blood coagulation. 1 The inherited bleeding disorder, hemophilia A, results from quantitative or qualitative deficiency of coagulation FVIII and affects 1 in 5000 males. There are approximately 17 000 patients with hemophilia in the United States, 80% of whom have hemophilia A. 2 This lifelong hemorrhagic diathesis is treated successfully with FVIII replacement either from plasma-derived sources or, for the last decade, primarily with recombinantly derived protein. 3 Plasma levels maintained above 2% can effectively prevent most severe hemorrhages, 4 so hemophilia A has been an attractive target for gene therapy applications.Recombinant FVIII (rFVIII) therapy has proved to be costly due to the expense of production, purification, and formulation. The manufacturing technology used has required specialized centralized production and distribution, limiting access to developing and third-world countries. 5 rFVIII still requires intravenous access for delivery due to limited bioavailability from other delivery routes. Regular prophylactic infusions of rFVIII can effectively prevent joint hemorrhages and prevent the development of hemophilic arthropathy. However, the cost and limited availability of rFVIII has prevented universal implementation of this treatment strategy. In addition, gene therapy applications for hemophilia A have been hampered by inadequate expression in vivo. 6,7 Several bioc...

The relations between normative developmental transitions, contextual influences, and life-stage outcomes such as academic achievement for African American males are examined. The integration of affective and cognitive processes are reviewed for preschool-and elementary-aged boys, and an empirical examination of vulnerable and resilient outcomes of 219 urban adolescent males is used to illustrate how structural conditions provide opportunities for vulnerable and successful academic outcomes. The results indicate that negative stereotyping and tracking from early experiences in educational settings influence African American males'scholastic achievement. By adolescence, given normative developmental experiences of social mobility and increased cognitive reasoning, experiences in their community as well as school contexts influences African American males' reactive or proactive coping strategies that influence vulnerability and resilience in school. Suggestions for future research and intervention efforts are offered.This article seeks to organize what we know about the affect of affective-and cognitive-linked developmental transitions on achievement outcomes and academic-associated processes, particularly for African American males. As an organizational strategy for accomplishing this task, the synthesis has three goals that also 608

No single theory so far proposed gives a wholly satisfactory account of the origin and maintenance of bird-song dialects. This failure is the consequence of a weak comparative literature that precludes careful comparisons among species or studies, and of the complexity of the issues involved. Complexity arises because dialects seem to bear upon a wide range of features in the life history of bird species. We give an account of the principal issues in bird-song dialects: evolution of vocal learning, experimental findings on song ontogeny, dialect descriptions, female and male reactions to differences in dialect, and population genetics and dispersal.We present a synthetic theory of the origin and maintenance of song dialects, one that accommodates most of the different systems reported in the literature. The few data available suggest that large, regional dialect populations are genetically differentiated; this pattern is correlated with reduced dispersal between dialects, assortative mating by females, and male-male exclusion. At the same time, “subdialects” may be formed within regional dialects. Subdialect clusters are usually small and may represent vocal mimicry among a few adjacent territorial males. The relative importance of genetic and social adaptation may contribute to the emergence of subdialects; their distinctiveness may be correlated with the degree of polygyny, for example. Thus, subdialect formation is linked to one theory of the evolution of repertoire size, but data are too fragmentary to examine this idea critically.

The research study investigated African American adolescent males' perceptions of their community resources and constraints. The participants were drawn from a sample of African American adolescents participating in a cross‐sectional longitudinal study in a large Southeastern American city. Research findings for two waves of data collection are reported [(8th, 9th, & 10th graders) and (10th, 11th, & 12th graders)]. The measures used were a combination of qualitative and quantitative instruments. The results from qualitative analyses indicated that adolescent males communicated with significant adults when confronted with negative experiences. Also, adolescent males reported awareness of negative neighborhood characteristics like drugs and violence. However, when students were asked about barriers that would keep them from reaching their life goals, over 2/3 of the sample had positive goal expectations. The measures used in the quantitative were subjected to reliability and validity checks. The instruments used had alphas above the .70 standard. The measures were The Black Male Experiences Measure, a revised‐Machismo Measure and a Revised‐Hassles Measure. The year 03 & 05 revised‐Machismo Measure (referred to as the Bravado Construct) was the dependent variable. Measures of daily hassles and negative/positive experiences were used to predict exaggerated bravado attitudes in males (F (4, 116) = 8.09, p > .001). The variance accounted for in the model was 19.64%. The implications of the study support beliefs that negative attitudes are learned and are associated with negative contextual experiences. Exaggerated male identity attitudes may be coping strategies for survival in high‐risk environments. Suggestions for prevention and intervention programs are given. © 1999 John Wiley & Sons, Inc.

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