Prevalence of cystic fibrosis mutations in Israeli Jews.

The aim of this study was to evaluate the screening policies of cystic fibrosis (CF) in the Jewish population. The prevalence of mutations that account for CF in Israel have been defined in the past by determining the frequency of CF mutations in affected individuals. This study is a population-based study and is, therefore, different from previous patient-based studies. We found that the CF mutations D1152H, W1089X, and 405 + IG-->A were present in some ethnic groups in which no CF patients carrying these mutations were reported. These facts necessitate a reevaluation of the screening policy regarding the ethnic groups in Israel. We studied 9,430 healthy Jewish Israeli individuals of 36 countries of origin. The prevalence of CF mutations was 1:19, 1:19, 1:28, and 1:42 for the Ashkenazi, Sephardi, North African, and Eastern Jews, respectively. CF mutations were identified in 374 (4.0%) individuals. These included 173 (46.3%) carriers of the W1282X mutation; 110 (29.4%) found to carry delF508; 23 (6.1%) who carried G542X; 22 (5.9%) who carried 3849 + 10Kb (C-->T; 20 (5.3%) who carried D1152H; 10 (2.7%) who carried N1303K; 11 (2.9%) who carried 405 + IG-->A; 4 (1.1%) who carried W1089X; and one (0.3%) who carried S549R. No carriers were detected for the 1717-1G-->A, G85E, and T360K mutations, which were tested for in 7,383, 1,558, and 41 individuals, respectively.

courtless, the Drosophila UBC7 homolog, is involved in male courtship behavior and spermatogenesis.

The courtless (col) mutation disrupts early steps of courtship behavior in Drosophila males, as well as the development of their sperm. Most of the homozygous col/col males (78%) do not court at all. Only 5% perform the entire ritual and copulate, yet these matings produce no progeny. The col gene maps to polytene chromosome band 47D. It encodes two proteins that differ in their carboxy termini and are the Drosophila homologs of the yeast ubiquitin-conjugating enzyme UBC7. The col mutation is caused by an insertion of a P element into the 3' UTR of the gene, which probably disrupts translational regulatory elements. As a consequence, the homozygous mutants exhibit a six- to sevenfold increase in the level of the COL protein. The col product is essential, and deletions that remove the col gene are lethal. During embryonic development col is expressed primarily in the CNS. Our results implicate the ubiquitin-mediated system in the development and function of the nervous system and in meiosis during spermatogenesis.

Shared HLA antigens in couples with 5 or more, compared to 3 or 4 consecutive recurrent miscarriages.

OBJECTIVE: Couples with consecutive recurrent miscarriages (CRM) are not generally believed to share more HLA antigens with their spouses than expected by chance. This paper attempted to determine the situation in patients with five or more miscarriages. METHODS: The number of shared HLA class I and class II antigens, HLA phenotype, and the ability to mount an antibody response in a large cohort of 425 couples with CRM was assessed, according to whether the patient had three or four, or five or more miscarriages. RESULTS: There was no significant difference in the frequency of shared HLA antigens in women with five or more miscarriages when compared to three or more miscarriages or control patients. No specific HLA antigen or phenotype was associated with CRM in male or female partners of either group. The number of shared antigens did not influence ability to develop anti paternal antibodies (APA). Moreover, HLA antigen sharing had no influence on the subsequent pregnancy after paternal leucocyte immunization. CONCLUSION: Class I and Class II HLA antigens are not diagnostic for immunologically mediated abortion, do not predict the ability to mount an antibody response, or the outcome of a subsequent pregnancy.

The prognostic value of anti-paternal antibodies and leukocyte immunizations on the proportion of live births in couples with consecutive recurrent miscarriages.

Anti-paternal antibodies directed towards paternal leukocytes have been used to predict the prognosis for the subsequent pregnancy in women with consecutive recurrent miscarriages (CRM) and also to determine if the patient has become immune after paternal leukocyte immunization. The predictive value is controversial, as these antibodies are not essential for pregnancy to develop, and only occur in a minority of parous women. This study tried to determine the predictive value of these antibodies when assessed separately for women with five or more abortions and compared to women with three or four abortions. The patients were assessed separately so that the higher live birth rate in the latter group would not obscure meaningful results in the former group with a poor prognosis. Antibody production, whether spontaneous, or induced by immunization, raised the live birth rate in primary and tertiary aborters with three, four, five or more abortions. Anti-paternal antibodies increased the proportion of live births from 18.5 to 53. 7% (P /= 5 CRM and 3-4 CRM respectively. Both immunization with paternal leukocytes per se and the ability to express anti-paternal antibodies were associated with an increased proportion of live births in the next pregnancy. Multivariate analysis showed that that the odds ratio for a live birth was approximately four times greater in women who were immunized and produced anti-paternal antibodies than in control patients. The lack of anti-paternal antibodies at initial testing could serve as a marker for the benefit of immunization with paternal leukocytes; the subsequent presence as a prognostic marker for the subsequent pregnancy.

The COP9 signalosome is essential for development of Drosophila melanogaster.

The COP9 signalosome (originally described as the COP9 complex) is an essential multi-subunit repressor of light-regulated development in plants [1] [2]. It has also been identified in mammals, though its role remains obscure [3] [4] [5]. This complex is similar to the regulatory lid of the proteasome and eIF3 [5] [9] [10] [11] [12] and several of its subunits are known to be involved in kinase signaling pathways [4] [6] [7] [8]. No proteins homologous to COP9 signalosome components were identified in the Saccharomyces cerevisiae genome, suggesting that the COP9 signalosome is specific for multi-cellular differentiation [13]. In order to reveal the developmental function of the COP9 signalosome in animals, we have isolated Drosophila melanogaster genes encoding eight subunits of the COP9 signalosome, and have shown by co-immunoprecipitation and gel-filtration analysis that these proteins are components of the Drosophila COP9 signalosome. Yeast two-hybrid assays indicated that several of these proteins interact, some through the PCI domain. Disruption of one of the subunits by either a P-element insertion or deletion of the gene caused lethality at the late larval or pupal stages. This lethality is probably a result of numerous pleiotropic effects. Our results indicate that the COP9 signalosome is conserved in invertebrates and that it has an essential role in animal development.

Overexpression Beadex mutations and loss-of-function heldup-a mutations in Drosophila affect the 3' regulatory and coding components, respectively, of the Dlmo gene.

LIM domains function as bridging modules between different members of multiprotein complexes. We report the cloning of a LIM-containing gene from Drosophila, termed Dlmo, which is highly homologous to the vertebrate LIM-only (LMO) genes. The 3' untranslated (UTR) of Dlmo contains multiple motifs implicated in negative post-transcriptional regulation, including AT-rich elements and Brd-like boxes. Dlmo resides in polytene band 17C1-2, where Beadex (Bx) and heldup-a (hdp-a) mutations map. We demonstrate that Bx mutations disrupt the 3'UTR of Dlmo, and thereby abrogate the putative negative control elements. This results in overexpression of Dlmo, which causes the wing scalloping that is typical of Bx mutants. We show that the erect wing phenotype of hdp-a results from disruption of the coding region of Dlmo. This provides molecular grounds for the suppression of the Bx phenotype by hdp-a mutations. Finally, we demonstrate phenotypic interaction between the LMO gene Dlmo, the LIM homeodomain gene apterous, and the Chip gene, which encodes a homolog of the vertebrate LIM-interacting protein NLI/Ldb1. We propose that in analogy to their vertebrate counterparts, these proteins form a DNA-binding complex that regulates wing development.

Metal ions suppress the abnormal taste behavior of the Drosophila mutant malvolio.

A mutation in the malvolio (mvl) gene affects taste behavior in Drosophila melanogaster. The malvolio gene encodes a protein (MVL) that exhibits homology to the mammalian natural resistance-associated macrophage proteins. It is also homologous to the Smf1 protein from Saccharomyces cerevisiae, which we have recently demonstrated to function as a Mn2+/Zn2+ transporter. We proposed that the Drosophila and mammalian proteins, like the yeast SMF1 gene product, are metal-ion transporters. To test this hypothesis, malvolio mutant flies were allowed to develop, from egg to adulthood, on a medium containing elevated concentrations of metal ions. Mutant flies that were reared in the presence of 10 mmol l-1 MnCl2 or FeCl2 developed into adults with recovered taste behavior. CaCl2 or MgCl2 had no effect on the mutant's taste perception. ZnCl2 inhibited the effect of MnCl2 when both ions were supplied together. Similar suppression of the abnormal taste behavior was observed when mvl mutants were fed MnCl2 or FeCl2 only at the adult stage. Furthermore, exposure of adult mutant flies to these ions in the testing plate for only 2 h was sufficient to restore normal taste behavior. The suppression of the defective taste behavior suggests that MVL functions as a Mn2+/Fe2+ transporter and that Mn2+ and/or Fe2+ are involved in the signal transduction of taste perception in Drosophila adults.

Behavioral analysis of Drosophila mutants displaying abnormal male courtship.

We describe six recessive autosomal male sterile mutations in Drosophila, generated by mobilization of single P-elements, exhibiting abnormal male courtship behavior. Detailed analysis of courtship behavior elicited by virgin wild type females indicated that five of the six mutants are affected in the early steps of courtship. The sixth mutant is blocked at the step of attempted copulation which occurs later in the courtship sequence. All of the mutants have normal olfactory responses and normal locomotor activity. No defect in the visual modality has been observed for the five mutants affected in the initiation of courtship. The mutant blocked at attempted copulation lacks the 'on' and 'off' transients, but this appears to be due to genetic background rather than the mutation itself. Abnormal spermatogenesis was observed in five of the mutants. Spermatogenic defects vary and include lesions in the proliferation of the germline, in meiosis, and in the differentiation and maturation of the spermatids into motile sperm.

The structure of protein phosphatase 2A is as highly conserved as that of protein phosphatase 1.

cDNA coding for protein phosphatase 2A (PP2A) has been isolated from Drosophila head and eye imaginal disc libraries. Drosophila PP2A mRNA is expressed throughout development, but is most abundant in the early embryo. The cDNA hybridises to a single site on the left arm of the second chromosome at position 28D2-4. The deduced amino acid sequence (309 residues) of Drosophila PP2A shows 94% identity with either rabbit PP2A alpha or PP2A beta, indicating that PP2A may be the most conserved of all known enzymes.

Immunization by paternal leukocytes for prevention of primary habitual abortion: results of a matched controlled trial.

Habitual abortion is a difficult clinical problem, as no cause can be found for abortion in over 50% of patients. At the habitual abortion clinic of the Sheba Medical Center, immunological activity is tested and patients who are considered suitable are offered immunopotentiation with paternal leukocytes. Patients are only treated if they have no other cause for habitual abortion, no lupus anticoagulant and no antipaternal complement-dependent antibodies (APCA). Immunization is thought to potentiate the maternal immune response to paternal antigens encountered on the trophoblast. The production of APCA antibody indicates that an immune response has occurred. Of the 156 patients so far immunized, 109 have developed these antibodies. To date, 79 of these 156 patients have become pregnant. Sixty-seven patients (with 3-12 miscarriages each) belong to the antibody-positive group. Sixty-four of the 89 subsequent pregnancies have been carried past their previous dates of abortion. Forty-seven live births have occurred. By contrast, 12 patients have been pregnant in the antibody-negative group, of the 16 subsequent pregnancies only 6 were successful. A control group is available for comparison. This consists of patients suitable for immunization, but not immunized. Of these patients, only 11 of 30 pregnancies have been carried to term.

Selection of patients with habitual abortion for paternal leucocyte immunization.

After potentiation of the immune response in habitual aborters 75-85% of subsequent pregnancies are claimed to result in healthy term infants. However, all publications to date have either been based on the authors concept of the immune processes involved or an attempt to demonstrate the efficacy of treatment either empirically or by matched trials. As immunization is coming into wider clinical use, it is necessary to determine which patients will benefit from this form of treatment. This paper presents our experience with paternal leucocyte immunization over the period 1985-1988. 207 patients were classified on a clinical basis and by immunological testing. 143 patients have been immunised, 129 pregnancies have occurred in 108 patients. The vast majority of our patients have recurrent missed abortions. Only six women habitually aborted live fetuses. Two had subsequent live births. Secondary aborters seem to do well in subsequent pregnancies, whether immunized or not. The patient most likely to benefit from immunization is the Primary missed aborter who does not possess antipaternal antibody (APCA), but is induced to produce APCA by immunization. Using these criteria, 75% success rates are observed in the subsequent pregnancy. This success rate is irrespective of HLA antigen sharing or in-vitro mixed lymphocyte reactivity.

The Drosophila Mutant tetanic Interacts with a Gene Complex Including the Structural Locus of K+ Channels and Shows Altered Dephosphorylation and Learning.

The tetanic (tta; X.-52.6) mutation has been isolated on the basis of its sensitivity to extradoses of the normal Shaker gene complex (ShC) where the K+ channel la is encoded. The mutant shows up to threefold elevation of the membrane bound protein phosphatase type 1 (PP1) activity in body extracts, probably due to reduced levels of the PP1 specific inhibitor 2 (I-2). By contrast, PP1 activity in the head is only half of the normal value. In addition, tta fails to perform normally in a negative reinforcement olfactory paradigm. The functional relationships between phosphorylation, K+ currents, phosphatase activity and modulation of synaptic activity during learning and memory are discussed in the light of their possible genetic links.

[Carrier detection and prenatal diagnosis in phenylketonuria, cystic fibrosis and adrenal hyperplasia use of molecular biology techniques].

With the advent of molecular biology techniques the prenatal diagnosis of many inherited diseases is now possible. In our Division of Transplantation Immunology we provide prenatal diagnosis for phenylketonuria (PKU), cystic fibrosis (CF) and congenital adrenal hyperplasia (CAH). In CF and PKU the chromosome carrying the disease gene is identified by the molecular probe, while in CAH it can also be determined by HLA phenotyping. Accurate diagnosis of a disease is dependent on the physical distance on the chromosome between the probe and the disease gene. Chorionic villous sampling allows evaluation of embryos at 9-10 weeks of gestation and also identification of carriers. DNA prepared from white blood cells of members of 4 families with CAH was digested with restriction endonucleases. Southern transfers were hybridized with the probe for 21-hydroxylase, and with 3 HLA probes mapped to both sides of the gene for 21-OH. In 2 families the embryo was found to be normal and in 2 diseased. Using the same techniques, but with probe and endonucleases specific for PKU, prenatal diagnosis was provided for 11 families with that condition. An embryo with PKU was found in each of 2 families, normal ones in 7, and in the remaining 2 families the testing was not informative. As of the present, 6 normal and 2 diseased children have been born, all as predicted. In 8 families with CF, DNA was examined with 5 probes mapped to both sides of the CF gene. Carriers and healthy sibs were identified, and in 1 family prenatal diagnosis was provided.

HLA-A11 is associated with poor prognosis in childhood acute lymphoblastic leukemia (ALL).

A possible association between HLA antigens, susceptibility or resistance to leukemia, and responsiveness to treatment has been studied in 144 patients with childhood acute lymphoblastic leukemia (ALL) and compared to other prognostic factors, i.e. white blood cell (WBC) counts, age at onset, sex, ethnic origin, and cell surface markers. All sequentially newly diagnosed children (97) comprised the group for the prospective study (PSG) and were followed for 6 years. The group included 37 patients classified as T-ALL, 41 as CALLA+, 27 as NULL, 12 as B and pre-B, and 27 unclassified patients, who were diagnosed before 1980. During the follow-up period, 45 patients of the PSG died. Forty-seven patients designated long-term survivors (LTS) have been followed 6-20 years after diagnosis, having completed a 3-5 year course of anti-leukemia therapy, and having remained disease free thereafter. High WBC counts at diagnosis and T-cell-surface markers were associated with poor prognosis, as were enthnic origin and specific HLA antigens. Thus, there was one (1) a significant increase in HLA-A30 and a decrease in HLA B-14 in the PSG Jewish patients; and (2) a complete absence of HLA-ALL in LTS while, in the PSG, 8 of 9 HLA-All-positive patients died during the follow-up period. This suggests that HLA-All is associated with poor prognosis in childhood ALL.

The protein phosphatases of Drosophila melanogaster and their inhibitors.

Protein phosphatases-1, 2A and 2B have been identified in membrane and soluble fractions of Drosophila melanogaster heads. Similarities between Drosophila and mammalian protein phosphatase-1 included specificity for the beta subunit of phosphorylase kinase, sensitivity to inhibitor-1 and inhibitor-2, inhibition by protamine, retention by heparin-Sepharose and selective interaction with membranes. In addition, an inactive form of protein phosphatase-1, termed protein phosphatase-1I, was detected in the soluble fraction that could be activated by preincubation with MgATP and mammalian glycogen synthase kinase-3. Inhibitor-2 partially purified from Drosophila had an identical molecular mass to its mammalian counterpart, and recombined with mammalian protein phosphatase-1 to form a hybrid protein phosphatase-1I. Similarities between Drosophila and mammalian protein phosphatase-2A included preferential dephosphorylation of the alpha subunit of phosphorylase kinase, insensitivity to inhibitors-1 and -2, activation by protamine, exclusion from heparin-Sepharose and apparent molecular mass. A Ca2+-dependent calmodulin-stimulated protein phosphatase (protein phosphatase-2B) that was inhibited by trifluoperazine was identified in the soluble fraction. The remarkable similarities between Drosophila protein phosphatases and their mammalian counterparts are indicative of strict phylogenetic conservation and demonstrate that the procedures used to classify mammalian protein phosphatases have a wider application. Characterisation of the Drosophila phosphatases will facilitate genetic analysis of dephosphorylation systems and their possible roles in neuronal and behavioural plasticity in Drosophila.

Antibodies to HIV in Israeli hemophiliacs: relationship between serological profile and disease development.

We studied 66 Israeli hemophiliacs for antibodies to HIV in blood samples collected between 1978 and 1985. By May 1985, 2 had AIDS, 2 had ARC, 4 had lymphadenopathy with some immunologic dysfunction, and 58 were asymptomatic. Antibodies to HIV were detected in 40 (60.6%) patients, including all 8 with disease. Presence of HIV antibodies was significantly associated with receipt of non-heat-treated commercial factor VIII concentrates (NHT fac VIII) between 1980 and 1983. Thirty-eight of 45 (84.44%) patients treated with NHT fac VIII developed antibodies to HIV, compared to 1 of 16 (6.25%) treated with cryoprecipitates and fresh plasma only. Of 40 seropositive patients, 1 (2.5%) had antibodies by 1980, 4 (10%) by 1982, 14 (35%) by 1983, 10 (25.0%) by 1984, and 11 (27.5%) by May 1985. The decline in the rate of seroconversion can be attributed to the replacement of NHT fac VIII concentrate with heat-inactivated factor VIII (HT fac VIII) concentrate by November 1983. As of January 1984 only HT fac VIII was administered. Twenty-nine multitransfused thalassemia patients as well as 20 healthy Israeli blood donors were seronegative to HIV. All 40 (100%) seropositive hemophiliacs had antibodies to viral env gene encoded gp120/gp160 antigens. Twenty-four (60.05%) also had antibodies to viral gag gene encoded p24 and/or p55 antigens. While antibodies to gp120/160 persisted during the follow-up time, a loss of antibodies to p24/55 was observed in 5 of 16 (31.25%) seropositive patients from whom multiple samples were available. gp120/160 positive, p24/55 negative hemophiliacs had significantly lower absolute T-helper cell counts and reversed Th/Ts ratios when compared to gp120/160 p24/55 seropositive patients. Four of the 16 (25.0%) asymptomatic gp120/160 positive, p24/55 negative patients developed overt disease within 15 months of the last blood collection. The data suggest that exposure to HIV antigens is widespread among hemophiliacs in Israel, and can be attributed to receipt of NHT fac VIII concentrates prior to 1984. Antibodies to gp120/160 are of the most important diagnostic value while loss of antibodies to p24/p55 may be of prognostic value.