Frequency of HLA-A alleles in the Syrian population genotyped by sequence-based typing.

HLA-A molecules are highly polymorphic. Their accurate typing at a high-resolution level is crucial for successful organ, bone marrow and cord blood transplantation. Furthermore, several HLA alleles have been involved in susceptibility to autoimmune diseases, allergies, cancers and inflammations. In order to determine common HLA-A alleles in Syria and their frequencies, sequence-based typing (SBT) was used to genotype HLA-A alleles at high resolution (four digit level) among one hundred and thirty randomly selected Syrian individuals. Exons 2, 3 and 4 of the HLA-A gene were amplified by PCR and sequenced. The sbt-engine software was used for allele assignment. Ambiguities were solved using group-specific sequencing primers (GSSPs). We could identify 32 different HLA-A alleles which were divided into 3 groups: high frequency (approximately 10%, A*01:01; A*24:02; A*03:01; A*02:01), moderate frequency (approximately 3%, such as A*02:05, A*31:01 and A*33:01), and low frequency (approximately 1%, such as A*02:11, A*29:01, A*02:02 and A*36:01). Homozygosity rate was higher than expected (11.5% vs. 7.15%). For high frequency alleles, our results show similarity to neighbouring countries. However, 15 alleles (such as A*02:04, A*02:06, A*02:11 and A*02:17) found in our cohort in low frequencies were never reported in some or all neighbouring countries. This is the first report on HLA-A allele frequencies in Syria. In spite of the relatively low number of tested subjects, our results revealed a high degree of diversity, with 32 different alleles, reflecting the high ethnic heterogeneity of the Syrian population. The identification of alleles rarely or never reported in neighbouring countries indicates a higher genetic diversity in Syria.

Drug resistance pulmonary tuberculosis in the Eastern Province of Saudi Arabia.

OBJECTIVE: To evaluate the prevalence and pattern of antituberculous drug resistance and patients with pulmonary tuberculosis in the Eastern Province and its impact on the tuberculosis control program. METHODS: Patients with pulmonary tuberculosis, proven by culture, admitted to Dammam Chest Hospital from November 1993 through May 1996 were reviewed. Patients who had at least one documented isolate of mycobacterium tuberculosis resistant to at least one standard anti-tuberculosis drug were identified. Medical records were reviewed and information was retrieved regarding age, sex, nationality, history of previous tuberculosis, human immune deficiency status, and results of direct smear and chest radiograph abnormalities. RESULTS: A total of 411 cases of culture positive pulmonary tuberculosis with drug susceptibility testing, were identified during the period mentioned, of these 123 (30%) were Saudi nationals and 228 (70%) were non-Saudis. Drug resistance to at least one drug was observed in 43 (10.5%) patients, resistance to isoniazid alone was observed in 25 (6%) patients, whereas resistance to rifampicin was noted in only one (0.2%) patient, and to streptomycin in 3 (1%) patients, multidrug resistance was observed in 11 (3%) patients. CONCLUSION: The study has shown that the overall drug resistance rate of 10.5% in the Eastern Province of Saudi Arabia is the lowest reported in the Kingdom, compared with Riyadh (13%), Taif (23%) and Gizan (44%). However, it appears to be similar to that reported in neighboring countries. In light of the study findings, and as per the recommendation of the World Health Organization, we suggest that a 4-drug regimen, as an initial treatment for pulmonary tuberculosis should be maintained, as resistance to isoniazid is still higher than the cut off value of 4%, and susceptibility testing for first line antituberculosis drugs should be carried out whenever possible. We also recommend applying stricter medical criteria for tuberculosis screening in newcomers, and for Saudi nationals, application of directly observed therapy should be a priority.

Functional analysis of six genes from chromosomes XIV and XV of Saccharomyces cerevisiae reveals YOR145c as an essential gene and YNL059c/ARP5 as a strain-dependent essential gene encoding nuclear proteins.

We report here basic functional analysis of strains deleted for six open reading frames (ORFs), YNL059c and YNL148c from chromosome XIV and YOR145c, YOR152c, YOR161c and YOR162c from chromosome XV of Saccharomyces cerevisiae. ORFs were replaced with the KanMX4 resistance marker using a long flanking homology PCR strategy in FY1679 and W303 diploid strains. Replacement cassettes were constructed in plasmid pUG7 and the cognate wild-type genes were recovered by gap repair. Sporulation and tetrad analysis revealed that deletion of YNL059c/ARP5 was lethal for vegetative growth in strain W303 and caused severe growth defects in strain FY1679 while YOR145c was essential for growth in both strains. Fusion of the green fluorescent protein (GFP) gene to the 3' ends of the YNL059c/ARP5 and YOR145c coding sequences created functional chimeric genes at the respective chromosomal loci. Both Arp5-GFP and Yor145-GFP localized to the nucleus, Yor145-GFP concentrating in the nucleolus. The vectors containing the deletion cassettes and the cognate wild-type genes, the oligonucleotides, and the deletant strains are available from the EUROFAN resource centre EUROSCARF (Frankfurt).

The Saccharomyces cerevisiae homologue of human Wiskott-Aldrich syndrome protein Las17p interacts with the Arp2/3 complex.

Yeast Las17 protein is homologous to the Wiskott-Aldrich Syndrome protein, which is implicated in severe immunodeficiency. Las17p/Bee1p has been shown to be important for actin patch assembly and actin polymerization. Here we show that Las17p interacts with the Arp2/3 complex. LAS17 is an allele-specific multicopy suppressor of ARP2 and ARP3 mutations; overexpression restores both actin patch organization and endocytosis defects in ARP2 temperature-sensitive (ts) cells. Six of seven ARP2 ts mutants and at least one ARP3 ts mutant are synthetically lethal with las17Delta ts confirming functional interaction with the Arp2/3 complex. Further characterization of las17Delta cells showed that receptor-mediated internalization of alpha factor by the Ste2 receptor is severely defective. The polarity of normal bipolar bud site selection is lost. Las17-gfp remains localized in cortical patches in vivo independently of polymerized actin and is required for the polarized localization of Arp2/3 as well as actin. Coimmunoprecipitation of Arp2p with Las17p indicates that Las17p interacts directly with the complex. Two hybrid results also suggest that Las17p interacts with actin, verprolin, Rvs167p and several other proteins including Src homology 3 (SH3) domain proteins, suggesting that Las17p may integrate signals from different regulatory cascades destined for the Arp2/3p complex and the actin cytoskeleton.

Analysis of a 35.6 kb region on the right arm of Saccharomyces cerevisiae chromosome XV.

We report the sequence of a 35,600 bp fragment covering the PET123 region on the right arm of chromosome XV from Saccharomyces cerevisiae. This region contains 19 possible open reading frames (ORFs) of which 16 are non-overlapping ORFs. Eight ORFs correspond to the SPP2, SMP3, PDR5, NFI1, PUP1, PET123 and MTR10 loci, described previously. Two ORFs correspond to yeast homologues of genes from other organisms: O3530 is a member of the large ribosomal subunit protein L13 family and O3560 (SME1 gene) is a 94-codon ORF and is a homologue of the mammalian SmE spliceosomal core protein. Three ORFs (O3513, O3521, O3548) present significant similarities to proteins of unknown function and three ORFs (O3510, O3536, O3545) lack homology to sequences within the databases screened.

Analysis of a 22,956 bp region on the right arm of Saccharomyces cerevisiae chromosome XV.

We present here the sequence analysis of a DNA fragment (cosmid pUOA1258) located on the right arm of chromosome XV. The 22,956 bp sequence reveals 14 open reading frames (ORFs) longer than 300 bp and the 201 bp RPS33 gene. Among the 14 large ORFs, two overlapping frames are likely to be non-expressed and one corresponds to the known GLN4 gene encoding glutaminyl-tRNA synthetase. Two ORFs, O3571 and O3620, encode putative transcriptional regulators with a Zn(2)-Cys(6) DNA binding domain characteristic of members of the GAL4 family. Among the nine remaining ORFs, five (O3568, O3575, O3590, O3615 and O3625) present significant similarity to proteins of unknown function and four (O3580, O3595, O3630 and O3635) lack homology to sequences present in the databases screened.

The Saccharomyces cerevisiae actin-related protein Arp2 is involved in the actin cytoskeleton.

Arp2p is an essential yeast actin-related protein. Disruption of the corresponding ARP2 gene leads to a terminal phenotype characterized by the presence of a single large bud. Thus, Arp2p may be important for a late stage of the cell cycle (Schwob, E., and R.P. Martin, 1992. Nature (Lond.). 355:179-182). We have localized Arp2p by indirect immunofluorescence. Specific peptide antibodies revealed punctate staining under the plasma membrane, which partially colocalizes with actin. Temperature-sensitive arp2 mutations were created by PCR mutagenesis and selected by an ade2/SUP11 sectoring screen. One temperature-sensitive mutant that was characterized, arp2-H330L, was osmosensitive and had an altered actin cytoskeleton at a nonpermissive temperature, suggesting a role of Arp2p in the actin cytoskeleton. Random budding patterns were observed in both haploid and diploid arp2-H330L mutant cells. Endocytosis, as judged by Lucifer yellow uptake, was severely reduced in the mutant, at all temperatures. In addition, genetic interaction was observed between temperature-sensitive alleles arp2-H330L and cdc10-1. CDC10 is a gene encoding a neck filament-associated protein that is necessary for polarized growth and cytokinesis. Overall, the immunolocalization, mutant phenotypes, and genetic interaction suggest that the Arp2 protein is an essential component of the actin cytoskeleton that is involved in membrane growth and polarity, as well as in endocytosis.