WIP regulates N-WASP-mediated actin polymerization and filopodium formation.

Induction of filopodia is dependent on activation of the small GTPase Cdc42 and on neural Wiskott-Aldrich-syndrome protein (N-WASP). Here we show that WASP-interacting protein (WIP) interacts directly with N-WASP and actin. WIP retards N-WASP/Cdc42-activated actin polymerization mediated by the Arp2/3 complex, and stabilizes actin filaments. Microinjection of WIP into NIH 3T3 fibroblasts induces filopodia; this is inhibited by microinjection of anti-N-WASP antibody. Microinjection of anti-WIP antibody inhibits induction of filopodia by bradykinin, by an active Cdc42 mutant (Cdc42(V12)) and by N-WASP. Our results indicate that WIP and N-WASP may act as a functional unit in filopodium formation, which is consistent with their role in actin-tail formation in cells infected with vaccinia virus or Shigella.

The human WASP-interacting protein, WIP, activates the cell polarity pathway in yeast.

WIP, the Wiskott-Aldrich syndrome protein-interacting protein, is a human protein involved in actin polymerization and redistribution in lymphoid cells. The mechanism by which WIP reorganizes actin cytoskeleton is unknown. WIP is similar to yeast verprolin, an actin- and myosin-interacting protein required for polarized morphogenesis. To determine whether WIP and verprolin are functional homologues, we analyzed the function of WIP in yeast. WIP suppresses the growth defects of VRP1 missense and null mutations as well as the defects in cytoskeletal organization and endocytosis observed in vrp1-1 cells. The ability of WIP to replace verprolin is dependent on its WH2 actin binding domain and a putative profilin binding domain. Immunofluorescence localization of WIP in yeast cells reveals a pattern consistent with its function at the cortical sites of growth. Thus, like verprolin, WIP functions in yeast to link the polarity development pathway and the actin cytoskeleton to generate cytoskeletal asymmetry. A role for WIP in cell polarity provides a framework for unifying, under a common paradigm, distinct molecular defects associated with immunodeficiencies like Wiskott-Aldrich syndrome.

Waltzing with WASP.

Wiskott-Aldrich syndrome (WAS) is an inherited immune deficiency that is marked by eczema, bleeding and recurrent infections. The lymphocytes and platelets of WAS patients display cytoskeletal abnormalities, and their T lymphocytes show a diminished proliferative response to stimulation through the T-cell receptor-CD3 complex (TCR-CD3). The product of the WAS gene, WAS protein (WASP), binds to the small GTPase Cdc42. Small GTPases of the Rho family are crucial for the regulation of the actin-based cytoskeleton. WASP and its relative NWASP might play an important role in regulating the actin cytoskeleton. Since both WASP and NWASP have the potential to bind to multiple proteins, they might serve as a hub to coordinate the redistribution of many cellular signals to the actin cytoskeleton. In this review, the authors discuss the possible role of WASP/NWASP and of the newly described protein WIP, which interacts with WASP and NWASP, in coupling signals from the T-cell receptor to the actin-based cytoskeleton.

Complete cDNA sequence of the HLA-DRB1*09012 allele.

Sequencing studies of HLA class II molecules have focused almost exclusively on exon 2. In this study the complete cDNA sequence of the DRB1*09012 allele is reported for the first time. This sequence was previously only partially published. In the DR9 antigen, two synonymous allelic variants (DRB1*09011 and 09012) were officially recognized, though it was later found that the first one contained an error and both sequences were, thus, identical.

C4d DNA sequences of two infrequent human allotypes (C4A13 and C4B12) and the presence of signal sequences enhancing recombination.

The DNA sequences of the polymorphic region (C4d) that belong to the infrequent complement C4 allotypes C4A13 and C4B12 have been obtained. In addition, C4A4 and C4B2 C4d sequences have been completed. C4A13 shows a new combination of amino acids at the following polymorphic positions: Asp1054, Pro1101 Cys1102, Leu1105, Asp1106, Asn1157, Ala1188, and Arg1191. These amino acids conform to the antigenic determinants Chido 1 and Rodgers 3; thus C4A13 is the only allele described thus far that carries both Ags. C4A13 and C4A4 carry the motif "ggctc*" (* means "deletion") at positions 14 to 19 in their intron 28; this motif had previously been reported only in C4B alleles. The C4B12 nucleotide sequence is analogous to C4B1b and C4B3 sequences, except for codon 1076, which is GCC in C4B1b and C4B3 and GGA in C4B12, which is coding for glycine in both cases. A recombination model for the generation of C4 alleles is formulated based on the analysis of these new sequences. One recombination would take place between positions 1157 and 1186 and would give rise to C4A13 and C4B5 or C4A3 (or C4A6) and C4B2; another one would occur between positions 1054 and 1076 and would generate C4A3 (or C4A6) and C4B12 or C4A2 and C4Bnew. Analysis of 1157 to 1186 and 1054 to 1076 fragments reveals the presence of putative sequence signals for recombination (similar to Escherichia coli chi recombination signal); the accumulation of such signals in fragments 1054 to 1076 supports the notion that a recombination hot spot for the C4 gene may exist and it also enhances new allele generation and intraspecies C4 gene homogenization.

Differential contribution of HLA-DR, DQ, and TAP2 alleles to systemic lupus erythematosus susceptibility in Spanish patients: role of TAP2*01 alleles in Ro autoantibody production.

OBJECTIVE: To study the influence MHC class II and TAP2 alleles exert on systemic lupus erythematosus (SLE) susceptibility and on the clinical and serological manifestations of the disease, in a cohort of Spanish patients. METHODS: HLA-DR serological typing and HLA-DQA, DQB, and TAP2 DNA sequence specific oligotyping, were carried out in 85 unrelated Spanish SLE patients and 186 healthy controls. Autoantibodies detection was carried out by indirect immunofluorescence and counter immunoelectrophoresis. RESULTS: Total SLE group: the frequency of HLA-DR3 and HLA-DQA1*0501 is significantly increased in this group (pc < 0.005, delta = 0.34 and pc < 0.005, delta = 0.45, respectively) although the highest delta value (delta = 0.87) is obtained when the TAP2*01 alleles are considered. No DQB allele shows significant deviation from the control group. Renal damage: it mainly occurs in HLA-DR3 patients (pc < 0.0005 and delta = 0.72). HLA-DQA1*0501 (p < 0.05, delta = 0.57 and DQB1*0201 (pc NS, delta = 0.56) are weaker susceptibility factors. Ro+ (but not LA) group: this autoantibody response is associated with TAP2*01 alleles in homozygosity (p < 0.05, delta = 0.81). R0/La+ group: it has a different genetic background as HLA-DQA1*0501 (delta = 1) and HLA-DQB1*0201 (delta = 1) are the main susceptibility factors. CONCLUSIONS: A differential association between HLA-DR, DQA1, and DQB1 alleles and SLE or its clinical and serological manifestations are found. Furthermore, the associations are different to the ones reported in other ethnic groups. Finally, TAP2*01 group of alleles are associated with the highest susceptibility to SLE (higher than HLA-DR3) and may influence Ro (but not La) autoantibodies production, whereas HLA-DQA1*0501 and DQB1*0201 mediates concomitant Ro and La productions.

Polymorphism of Mhc-DRB alleles in Cercopithecus aethiops (green monkey): generation and functionality.

DRB genes have been studied for the first time in green monkeys (Cercopithecus aethiops). Eleven new DRB alleles (exon 2, exon 3) have been obtained and sequenced from cDNA. A limited number of lineages have been identified: DRB1*03 (4 alleles), DRB1*07 (3 alleles), DRB5 (1 allele), DRB*w6 (1 allele), and DRB*w7 (2 alleles). The existence of Ceae-DRB1 duplications is supported by the finding of 3 DRB1 alleles in 3 different individuals. Ceae-DRB1*0701 may be non-functional because it bears serine at position 82, which hinders molecule surface expression in mice; the allele is only found in Ceae-DRB duplicated haplotypes. Base changes in cDNA Ceae-DRB alleles are consistent with the generation of polymorphism by point mutations or short segment exchanges between alleles. The eleven green monkey DRB alleles meet the requirements for functionality as antigen-presenting molecules (perhaps, excluding DRB1*0701), since: 1) they have been isolated from cDNA and do not present deletions, insertions or stop codons: 2) structural motifs necessary for a correct folding of the molecule, for the formation of DR/DR dimers and for CD4 interactions are conserved, and 3) the number of non-synonymous substitutions is higher than the number of synonymous substitutions in the peptide binding region (PBR), while the contrary holds true for the non-PBR region.

Complete nucleotide sequence of the HLA-DRB1 *1302 allele.

Sequencing studies of HLA class II molecules have focused almost exclusively on its exon 2. The complete cDNA sequence of the DRB*1302 allele is reported here for the first time. A conservative polymorphism was detected outside the exon 2 at residue 206; these changes can only be recorded if complete cDNA sequences are studied, and may be of value in drawing evolutive inferences.

Diploid expression of human leukocyte antigen class I and class II molecules on spermatozoa and their cyclic inverse correlation with inhibin concentration.

A diploid expression of class I and class II human leukocyte antigens (HLA) has been found in purified spermatozoa by using double fluorescence labeling cytofluorometry and relevant monoclonal antibodies; this expression has been confirmed for the first time by the analysis of specific HLA mRNA and metabolic 35S labeling followed by immunoprecipitation, which demonstrates an active ongoing translation of HLA proteins in germinal cells. Long-living mRNA coming from diploid germinal cells may be translated to HLA molecules in spermatozoa. This translation is controlled (or at least inversely correlated) by a testicular hormone (inhibin) in a cyclic fashion. Remarkably, serum levels of inhibin, synthesized by Leydig and Sertoli cells, follow a 12- to 13-day cycle, with a peak level at Day 6; this is probably controlled by FSH (not cyclic in males) and other testicular and/or unknown hormones. Peak levels of inhibin concur with the lower density and percentage of spermatozoa expressing both HLA class I and II molecules (close to 3% by cytofluorometry); lowest levels of inhibin coincide with the highest numbers (35-40%) of spermatozoa positive for both HLA molecules and a higher surface density. These observations could put to an end a disconcerting and long-lasting controversy on the expression/non-expression of HLA antigens on spermatozoa. The possibility that HLA-bearing spermatozoa are more capacitated for fertilization than those that do not bear HLA, and the implications of our results on male fertility control are also discussed.

HLA DR and DQ polymorphism in Ashkenazi and non-Ashkenazi Jews: comparison with other Mediterraneans.

HLA-DR and DQ alleles have been detected by DNA typing in Ashkenazi and non-Ashkenazi Jews from Israel. Allele frequencies, characteristic DR/DQ linkage disequilibria, population distances and their corresponding dendrogram by using the Neighbor-Joining method were used to study relatedness between Jewish and other Mediterranean and non Mediterranean populations. Closest relatedness is observed between Ashkenazi and non-Ashkenazi Jews, and, in decreasing order, also with Algerians, Spaniards (including Spanish-Basques), French and Italians. Also, particular characteristic Central European alleles are observed in Ashkenazi Jews and Mediterranean/African alleles in non-Ashkenazi Jews. This is consistent with historical data, Jews being an ancient Mediterranean population, who have had a certain degree of admixture with their 2000-3000 years old neighbors in spite of cultural and religious traditions which have preserved identity outside Israel.

The contribution of the HLA-A, -B, -C and -DR, -DQ DNA typing to the study of the origins of Spaniards and Basques.

The high polymorphism of the HLA system has been used as a powerful genetic tool to single out individuals and populations. By studying characteristic allele frequencies and extended HLA haplotypes in different populations, it is possible to identify ethnic groups and establish the genetic relationships among them. In the present study, HLA-A, -B, -C, -DR and -DQ typing at the serological/antigenic and the DNA level has been used for the first time to assign specific HLA frequencies and haplotypes to Spaniards and Basques and compare them with frequencies in other populations, particularly with North Africans. Allelic frequencies do not significantly differ between Spaniards and Basques. HLA genetic distances and their respective dendrogram together with the results on complete HLA haplotypes place Basques and Spaniards closer to paleo-North African populations than to other Europeans. This goes in favour of the Basques being a relative genetic isolate coming from the primitive Iberian/paleo-North African people. In addition, a tentative assignment of the most common Spanish HLA haplotypes to the different people who populated Iberia according to historical records has been done.

Susceptibility to insulin-dependent diabetes mellitus and short cytoplasmic ATP-binding domain TAP2*01 alleles.

TAP2 genes are placed within the HLA complex, have limited genetic variability and encode two main groups of peptide transporter proteins, the so-called TAP2*01 alleles, with a short ATP-binding domain, and the TAP2*0201 allele with a long domain. These transporters carry antigenic peptides from cytoplasm across the endoplasmic reticulum membrane to release them into nascent HLA class I molecules, which will then travel towards the plasma membrane. The shorter TAP2*01 alleles are present in 99% of diabetics and 90% of controls; these alleles may add slight, although significant and independent, susceptibility to diabetes, particularly in subjects carrying non-Asp 57 at beta DQ. Moreover, this increased susceptibility is not due to linkage disequilibrium with other HLA markers (i.e.: DR4), which does not exist in our Spanish population.