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PubMed1994

Castleman's disease. Differences in follicular dendritic network in the hyaline vascular and plasma cell variants.

Twenty-seven cases of the hyaline vascular variant and 10 cases of the plasma cell variant of Castleman's disease were studied with the paraffin resistant monoclonal antibodies Ki-FDC1p and/or Ki-M4p against follicular dendritic cells. Studies with the monoclonal antibody Ki-M9, for the detection of sinus lining cells, were also performed on the available frozen tissue in four cases of the hyaline vascular variant. In nine of the 10 plasma cell variant cases, the predominant type of follicular dendritic cell network was similar to that seen in normal or reactive germinal centres. In contrast, the hyaline vascular variant demonstrated either an expanded, disrupted, follicular dendritic cell network (10 cases) or multiple tight collections of follicular dendritic cells (16 cases). Sinus lining cells were not detected in the four cases studied. The difference in the predominant type of dendritic meshwork is an additional distinguishing feature to separate the plasma cell and hyaline vascular variants of Castleman's disease. The patterns of dendritic network seen in the hyaline vascular type, together with the absence of sinus lining cells, appear to favour the hamartoma theory proposed for this variant.

Castleman Disease↗

PubMed2006

Histopathological features of breast cancer in carriers of ATM gene variants.

AIMS: Germline variants in the ataxia telangiectasia mutated (ATM) gene have been implicated in increased breast cancer risk. The aim of this study was to determine whether the histopathology of breast cancers occurring in ATM variant carriers is distinctive or resembles the described BRCA1 mutation-associated phenotype. METHODS: The histopathological features of breast cancers occurring in ATM variant carriers from multiple-case breast cancer families were compared with matched controls. The test group included 21 cases of in situ and/or invasive cancer from carriers of either the IVS10-6T-->G, 2424V-->G or 1420L-->F ATM variants in the absence of BRCA1 or BRCA2 mutations. An additional four invasive cancers from carriers of a pathogenic BRCA1 mutation in the context of a familial ATM variant were also examined. RESULTS: The histopathology of breast cancers in ATM variant-only carriers was not significantly different from controls and known features of BRCA1 mutation-associated cancer were rarely seen. In contrast, these features were prominent in the small group of cases with a pathogenic BRCA1 mutation. CONCLUSIONS: Breast cancer occurring in carriers of ATM variants is not associated with distinctive histopathological features and does not resemble the tumour phenotype commonly observed in BRCA1 mutation carriers.

Adult↗

PubMed1990

Two of the three genetic variants of goat alpha s1-casein which are synthesized at a reduced level have an internal deletion possibly due to altered RNA splicing.

This paper describes the elucidation of the primary structure of the three genetic variants of goat alpha s1-casein, alpha s1-Cn D, E and F, which have been found to be associated with reduced amounts of alpha s1-casein in milk. Variant E has the same electrophoretic mobility as variant B, but differs from the latter by the substitutions of Arg for Lys and of Thr for Ala at positions 100 and 195. A genetically controlled event which does not affect the amino acid sequence of this variant might be responsible for its lower rate of synthesis compared to that of alpha s1-casein B. The deletion of 11 amino acids at positions 59-69 and of 37 amino acids at positions 59-95 in variant B leads to variants D and F. In both cases the deletions, which start at the same position of the polypeptide chain, include the major phosphorylation site of the protein. On the basis of sequence data for casein genes and cDNAs, it was concluded that the deletions occurring in the D and F variants are due to the exclusion of one and several exons, respectively. The observed deletions in the proteins could thus be the consequence of splice site mutations which would induce altered RNA processing and hence reduce the rate of synthesis of the casein.

Amino Acid Sequence↗

PubMed1987

Frequency and distribution of rare electrophoretic mobility variants in a population of human newborns in Ann Arbor, Michigan.

We have summarized the frequency and distribution of the rare variants encountered during the screening of 258,815 allele products, the products of 51 different loci, in 3242 predominantly Caucasian (88%) newborns. Seventy-nine different rare variants, representing 187 occurrences, were identified. Almost 60% (46 of 79) of the rare variants occurred as singletons while another 20% were seen in two unrelated individuals. No rare variants were detected at 18 loci while no variants, either rare or polymorphic, were detected at 14 loci. More rare variants were identified at loci that were classified as polymorphic and also at loci where the gene products exist as a monomer. A positive relationship was observed between variant frequency, either classes or copies, and subunit molecular mass.

Alleles↗

PubMed2006

The European-specific mitochondrial cluster J/T could confer an increased risk of insulin-resistance and type 2 diabetes: an analysis of the m.4216T > C and m.4917A > G variants.

The aims of this study were to investigate the contributions of the mitochondrial DNA m.4216T > C and m.4917A > G variants, and also of the European-specific mitochondrial cluster J/T, to the development of type 2 diabetes mellitus in Caucasian-Brazilian patients from Southern Brazil. We analyzed 347 type 2 diabetes patients and 350 control subjects. Variant frequencies in patients and control subjects were compared using chi2 tests or odds ratio. We also compared clinical and laboratory characteristics among patients with and without the variants. We found that the frequencies of the m.4216T > C and m.4917A > G variants are higher in diabetic patients than in control subjects. Moreover, haplogroups J (partially defined by the presence of the m.4216T > C variant only) and T (partially defined by the presence of both m.4216T > C and m.4917A > G variants) are more frequent in the type 2 diabetic group than in the control group. Patients belonging to the cluster J/T are more insulin resistant than patients of other haplogroups. In conclusion, our results indicate the association of the cluster J/T (as suggested by analyses of the m.4216T > C and m.4917A > G variants) with insulin resistance and type 2 diabetes.

Brazil↗

PubMed2006

Developmentally-regulated expression of tissue-specific splice variant of rat vesicular glutamate transporter 1 in retina and pineal gland.

Three distinct subtypes of vesicular glutamate transporters (VGLUTs) have been identified to date that are expressed basically in a cell type-specific manner. We have found a splice variant of VGLUT1 mRNA that is expressed almost exclusively in photosensitive tissues, i.e. the retina and the pineal gland. The variant mRNA, termed VGLUT1v, contains an additional 75 base pair sequence derived from part of a second intron (designated as exon IIa) between exons 2 and 3. The variant accounted for approximately 70% and 25%of VGLUT1 mRNA in the adult retina and pineal gland, respectively. The expression of VGLUT1v was developmentally regulated in both tissues. Organ culture showed that expression of the variant in the retina increased in association with the development of rod cells, suggesting that VGLUT1v is expressed in rod cells. In situ hybridization with variant-specific probes showed expression of VGLUT1v in the inner segment layer of photoreceptor cells. On the other hand, variant expression did not parallel the development of rhodopsin-positive cells in the pineal gland. As rod cells and pinealocytes are known to release glutamate continuously at ribbon synapses, it is possible that the variant has some functional advantage over the wild-type transporter in such a specialized manner of glutamate release.

Alternative Splicing↗

PubMed1999

T1762/A1764 variants of the basal core promoter of hepatitis B virus; serological and clinical correlations in Chinese patients.

BACKGROUND: A double variant in the basal core promoter, converting nucleotide 1762 from A to T (T1762) and nucleotide 1764 from G to A (A[764), has been described in patients with chronic hepatitis B infection. Its prevalence and significance in Chinese chronic HBV carriers are unknown. METHODS: We studied 177 Chinese patients with chronic HBV infection (chronic hepatitis/asymptomatic: 89/88; hepatitis B e antigen positive/negative: 84/93). The double variant was detected by mismatched polymerase chain reaction and restriction fragment length polymorphism analysis. The reliability of this method was verified by sequencing in 41 serum samples with 100% matching. RESULTS: The double variant T1762/ A1764 was found in 52 of 89 patients with chronic hepatitis, but in only 6 of 59 asymptomatic carriers (p<0.001). The prevalence was significantly lower in hepatitis B e antigen positive patients (23/84) than in hepatitis B e antigen negative patients (35/64) (p<0.005). Precore variant, A1896 was detected in 40 individuals; 31 of them suffered from chronic hepatitis and 9 were asymptomatic (p<0.001). A combination of both variants T1762/A1764 and A1896 was seen in 3 of 59 asymptomatic and 22 of 89 patients with chronic hepatitis (p<0.005). CONCLUSIONS: Mismatched polymerase chain reaction with restriction fragment length polymorphism provides a reliable, easy and fast method for detection of the presence of the T1762/A1764 variant. In Chinese chronic hepatitis B carriers, T1762/A1764 variant was associated with both active liver disease and hepatitis B e antigen negativity.

Adult↗

PubMed1999

Emergence of a laccase-positive variant of Azospirillum lipoferum occurs via a two-step phenotypic switching process.

Two variants have been isolated from the wild-type Azospirillum lipoferum strain 4B. The first variant, 4V(I), spontaneously emerged from the wild-type at frequencies in the order of 10(-4) to 10(-3) per cell generation. Compared to the wild-type, the 4V(I) variant gained (production of a carotenoid-like pigment, assimilation of certain carbohydrates) and lost (swimming motility, reduction of triphenyl tetrazolium chloride, acid production from certain sugars) apparently unrelated phenotypic characteristics. Only from the 4V(I) variant, a second atypical stable form, variant 4V(II), which acquired laccase activity and ability to produce melanin, appeared under very specific conditions, namely growth at extremely low oxygen concentrations. Neither of the variants was able to revert to the parental phenotype. The results suggest that atypical non-motile laccase-positive isolates of A. lipoferum that are found in the rice rhizosphere originate from wild-type (motile, laccase-negative) cells via a two-step phenotypic switching event, a non-motile laccase-negative variant being an intermediate phase.

Azospirillum↗

PubMed2006

Variants of beta-microglobulin cleaved at lysine-58 retain the main conformational features of the native protein but are more conformationally heterogeneous and unstable at physiological temperature.

Cleavage of the small amyloidogenic protein beta2-microglobulin after lysine-58 renders it more prone to unfolding and aggregation. This is important for dialysis-related beta2-microglobulin amyloidosis, since elevated levels of cleaved beta2-microglobulin may be found in the circulation of dialysis patients. However, the solution structures of these cleaved beta2-microglobulin variants have not yet been assessed using single-residue techniques. We here use such methods to examine beta2-microglobulin cleaved after lysine-58 and the further processed variant (found in vivo) from which lysine-58 is removed. We find that the solution stability of both variants, especially of beta2-microglobulin from which lysine-58 is removed, is much reduced compared to wild-type beta2-microglobulin and is strongly dependent on temperature and protein concentration. 1H-NMR spectroscopy and amide hydrogen (1H/2H) exchange monitored by MS show that the overall three-dimensional structure of the variants is similar to that of wild-type beta2-microglobulin at subphysiological temperatures. However, deviations do occur, especially in the arrangement of the B, D and E beta-strands close to the D-E loop cleavage site at lysine-58, and the experiments suggest conformational heterogeneity of the two variants. Two-dimensional NMR spectroscopy indicates that this heterogeneity involves an equilibrium between the native-like fold and at least one conformational intermediate resembling intermediates found in other structurally altered beta2-microglobulin molecules. This is the first single-residue resolution study of a specific beta2-microglobulin variant that has been found circulating in dialysis patients. The instability and conformational heterogeneity of this variant suggest its involvement in beta2-microglobulin amyloidogenicity in vivo.

Electrophoresis, Capillary↗

PubMed2005

Identification of four novel exon 5 splice variants of the mouse mu-opioid receptor gene: functional consequences of C-terminal splicing.

The rat mu-opioid receptor clone in which novel exon 5 was found in the place of exon 4 (MOR-1B) was one of the first MOR-1 variants described. We now have identified the mouse homolog of the rat MOR-1B as well as four additional variants derived from splicing from exon 3 into different sites within exon 5. The sequences of all of the variants were identical except for the intracellular tip of the C terminus encoded by exon 5, where each variant predicted a unique amino acid sequence ranging from 2 to 39 amino acids. All of the mMOR-1B variants were selective for mu-opioids in receptor-binding assays, as anticipated, because they all have identical binding pockets defined by the transmembrane domains. However, the relative potency and efficacy of mu-agonists to each other varied from variant to variant in guanosine 5'-O-(3-[35S]thio)triphosphate-binding studies, as shown by morphine-6beta-glucuronide, which was the most efficacious agent against mouse MOR-1B1 (mMOR-1B1) and the least efficacious agent against mMOR-1B2. mMOR-1B4 was quite unusual. Although mMOR-1B4 was mu-selective in receptor-binding studies and antagonists labeled mMOR-1B4 well, the binding affinities of most of the mu-agonists were far lower than those seen with mMOR-1, suggesting that the 39 amino acids at the C terminus of mMOR-1B4 influences the conformation of the receptor and its ligand recognition site itself either directly or through its interactions with other proteins. In conclusion, alterations in the amino acid sequence of the C terminus do not alter the mu-specificity of the receptor but they can influence the binding characteristics, efficacy, and potency of mu-opioids.

Animals↗

PubMed2001

Identification and functional characterization of a new CYP2C9 variant (CYP2C9*5) expressed among African Americans.

CYP2C9 is a polymorphic gene for which there are four known allelic variants; CYP2C9*1, CYP2C9*2, CYP2C9*3, and CYP2C9*4. In the present study, DNA from 140 European Americans and 120 African Americans was examined by single-strand conformational polymorphism and restriction fragment length polymorphism analyses, resulting in the identification of a new CYP2C9 variant, CYP2C9*5. This variant is derived from a C1080G transversion in exon 7 of CYP2C9 that leads to an Asp360Glu substitution in the encoded protein. The CYP2C9*5 variant was found to be expressed only in African Americans, such that approximately 3% of this population carries the CYP2C9*5 allele. The variant was expressed in, and purified from, insect cells infected with a recombinant baculovirus. Comparative kinetic studies using the purified wild-type protein CYP2C9*1; the Ile359Leu variant, CYP2C9*3; and the Asp360Glu variant, CYP2C9*5 were carried out using (S)-warfarin, diclofenac, and lauric acid as substrates. The major effect of the Asp360Glu mutation was to increase the K(m) value relative to that of CYP2C9*1 for all three substrates: 12-fold higher for (S)-warfarin 7-hydroxylation, 5-fold higher for the 4'-hydroxylation of diclofenac, and 3-fold higher for the omega-1 hydroxylation of lauric acid. V(max) values differed less than K(m) values between the CYP2C9*1 and CYP2C9*5 proteins. In vitro intrinsic clearances for CYP2C9*5, calculated as the ratio of V(max)/K(m), ranged from 8 to 18% of CYP2C9*1 values. The corresponding ratio for CYP2C9*3 was 4 to 13%. Accordingly, the in vitro data suggest that carriers of the CYP2C9*5 allele would eliminate CYP2C9 substrates at slower rates relative to persons expressing the wild-type protein.

Black or African American↗

PubMed2002

Metabotropic glutamate receptor 1 internalization induced by muscarinic acetylcholine receptor activation: differential dependency of internalization of splice variants on nonvisual arrestins.

In this study, we characterized the glutamate- or second-messenger kinase-dependent internalization of the rat metabotropic glutamate receptor 1 (mGluR1) splice variants 1a, 1b, and 1c, and assessed the arrestin and dynamin dependence of these processes. To facilitate this we inserted a hemagglutinin epitope tag in the extracellular N-terminal domain of the splice variants. Quantification of glutamate-induced mGluR1 splice variant internalization provided by enzyme-linked immunosorbent assay and confirmed by immunofluorescent microscopy indicated that each splice variant underwent rapid internalization, which was strongly inhibited by coexpression of dominant-negative mutant (DNM) arrestin or dynamin. In addition glutamate-induced rapid translocation of arrestin-2-green fluorescent protein (GFP) or arrestin-3-GFP from cytosol to membrane was observed in cells expressing mGluR1 splice variants. Glutamate-induced internalization of mGluR1a and mGluR1c was partially blocked by a selective inhibitor of protein kinase C (PKC), 2-[1-(3-dimethylamino-propyl)indol-3-yl]-3-(1H-indol-3-yl)maleimide (GF 109203X), whereas mGluR1b internalization was not significantly affected by this inhibitor. Similarly, inositol phosphate production after glutamate-induced activation of mGluR1a and mGluR1c was increased after PKC inhibition, whereas glutamate-induced mGluR1b stimulation was unaffected. Activation by carbachol of endogenously expressed M(1) muscarinic receptors in human embryonic kidney 293 cells, induced the internalization of mGluR1 splice variants, which was partially blocked by pretreatment with inhibitors of either PKC or Ca(2+) calmodulin-dependent kinase II (CaMKII). Expression of DNM-arrestin with mGluR1a or 1c strongly inhibited carbachol-induced internalization. However, coexpression of DNM-arrestin with mGluR1b was less effective in reducing carbachol-induced receptor internalization. In addition, arrestin-2-GFP or arrestin-3-GFP underwent significant carbachol-induced translocation from cytosol to membrane in cells coexpressing mGluR1a or 1c but not in cells coexpressing mGluR1b. This study demonstrates that the internalization of mGluR1 splice variants is subject to PKC and CaMKII regulation. In addition, regulation by these kinases confers differential arrestin dependence.

Alternative Splicing↗

PubMed2004

Identification of novel alternative splice variants of human constitutive androstane receptor and characterization of their expression in the liver.

Human constitutive androstane (or active) receptor (hCAR), a member of the nuclear receptor superfamily NR1I3, regulates the expression of several genes that are mainly involved in the metabolism of endogenous and xenobiotic compounds (e.g., CYP2B6, CYP3A4, and UGT1A1). We found four novel splice variants in the ligand-binding domain (LBD) of hCAR (NCBI reference sequence, NM_005122; designated SV0 herein). The variants designated SV1 and SV2 contained in-frame 12- and 15-base pair (bp) insertions, respectively. SV3 carried both of the insertions, and SV4 contained an in-frame 117-bp deletion. The insertion site of SV1 is located in the alpha6 helix of hCAR LBD, which makes up the ligand-binding cavity, and that of SV2 is located in the highly conserved loop between helices alpha8 and alpha9. SYBR Green real-time reverse transcription-polymerase chain reaction analysis of each splice variant revealed that the hepatic expression of SV2 was almost comparable with that of SV0 (approximately 40%), whereas other variants accounted for 6 to 10% of the total hCAR transcripts. In the reporter gene assays employing the phenobarbital-responsible enhancer module (PBREM) from CYP2B6 and UGT1A1 genes, the splice variants, except for SV1, were inactive, whereas SV1 transactivated the CYP2B6 PBREM but not the UGT1A1 PBREM reporter. A nuclear translocation assay in rat hepatocytes revealed that all the splice variants lack the responsiveness toward phenobarbital and 6-(4-chloropheny-l)imidazo[2,1-b][1,3]thiazole-5-carbaldehyde O-(3,4-dichlorobenzyl)oxime (CITCO) in terms of the ligand-dependent nuclear translocation. Further characterization, such as the identification of specific ligands, will help elucidate physiological implication of these hCAR splice variants.

Alternative Splicing↗

PubMed2026

Proteomics identify disease-associated variants in patients with rare diseases undiagnosed after genome sequencing.

Despite the introduction of genome sequencing (GS) for rare disease diagnostics, a genetic cause is not identified in most patients. Here, we explored the potential of proteomics to improve the diagnostic yield in 424 patients with rare diseases from the 100,000 Genomes Project (100kGP) without a genetic diagnosis. Serum proteomic profiling was performed using the Olink Explore 1536 assay (N = 1463 proteins). For 13 patients without genetic diagnoses, detection of lower serum protein "outliers" (z-score < -2) led to confirmed genetic diagnoses by resolving variants of uncertain significance or prioritizing genes for targeted GS reanalysis. For 23 additional patients without genetic diagnoses (64% of findings), we identified candidate gene-disease links and variants through convergent evidence from lower protein outliers and variants ranked through the variant prioritization tool Exomiser. For example, we identified a candidate heterozygous missense variant [Genome Aggregation Database (gnomAD) minor allele frequency = 0.006%] in tyrosine kinase with immunoglobulin-like and epidermal growth factor homology domains 1 (TIE1) that was only present in a patient with lower TIE1 serum abundance (z-score = -5.12) and their father, both of whom were affected by the same monogenic cardiac disorder, but in no other individuals from the 100kGP. Missense (52.5%) and splice region (27.5%) variants accounted for most diagnostic or candidate variants prioritized. This proof-of-principle study demonstrated that serum proteomics can support rare disease diagnosis and identify disease-causing genes in patients undiagnosed after GS, although successful implementation will likely depend on tissue specificity of protein expression, detectability in blood, proteomic platform coverage, and sensitivity.

Humans↗

PubMed2006

Isolation and characterization of brewer's yeast variants with improved fermentation performance under high-gravity conditions.

To save energy, space, and time, today's breweries make use of high-gravity brewing in which concentrated medium (wort) is fermented, resulting in a product with higher ethanol content. After fermentation, the product is diluted to obtain beer with the desired alcohol content. While economically desirable, the use of wort with an even higher sugar concentration is limited by the inability of brewer's yeast (Saccharomyces pastorianus) to efficiently ferment such concentrated medium. Here, we describe a successful strategy to obtain yeast variants with significantly improved fermentation capacity under high-gravity conditions. We isolated better-performing variants of the industrial lager strain CMBS33 by subjecting a pool of UV-induced variants to consecutive rounds of fermentation in very-high-gravity wort (>22 degrees Plato). Two variants (GT336 and GT344) showing faster fermentation rates and/or more-complete attenuation as well as improved viability under high ethanol conditions were identified. The variants displayed the same advantages in a pilot-scale stirred fermenter under high-gravity conditions at 11 degrees C. Microarray analysis identified several genes whose altered expression may be responsible for the superior performance of the variants. The role of some of these candidate genes was confirmed by genetic transformation. Our study shows that proper selection conditions allow the isolation of variants of commercial brewer's yeast with superior fermentation characteristics. Moreover, it is the first study to identify genes that affect fermentation performance under high-gravity conditions. The results are of interest to the beer and bioethanol industries, where the use of more-concentrated medium is economically advantageous.

Beer↗

PubMed2003

Differences in clinical manifestations and hematological and serological responses after experimental infection with genetic variants of Anaplasma phagocytophilum in sheep.

Five-month-old lambs were experimentally infected with two 16S rRNA genetic variants of Anaplasma phagocytophilum, variants 1 (GenBank accession no. M73220) and 2 (GenBank accession no. AF336220). Additional sequencing of the groESL heat shock operon gene indicated that these variants differ in three nucleotides at positions 782, 824, and 890. The variants were obtained by blood sampling of A. phagocytophilum-infected lambs from one sheep flock in Norway and were stored at -70 degrees C with 10% dimethyl sulfoxide as a cryoprotectant before being inoculated intravenously into susceptible lambs. The infectious blood contained, per ml, approximately 0.5 x 10(6) neutrophils infected with either of the variants. Six weeks after the primary inoculation, the lambs were challenged with the same infectious dose of the heterologous variant. The results of the study indicate a marked difference in clinical manifestation, neutropenia, antibody response, and cross-protection after experimental infection with the two variants of A. phagocytophilum.

Anaplasma phagocytophilum↗

PubMed1994

Specific genetic variants of Actinobacillus actinomycetemcomitans correlate with disease and health in a regional population of families with localized juvenile periodontitis.

A geographically homogeneous population of 83 subjects, from 21 families with localized juvenile periodontitis (LJP), and 35 healthy control subjects was monitored, over a 5-year period, for the presence of the periodontal pathogen Actinobacillus actinomycetemcomitans. Restriction fragment length polymorphism (RFLP) analysis was used to monitor the distribution of genetic variants of this bacterium in LJP-susceptible subjects that converted from a healthy to a diseased periodontal status. A. actinomycetemcomitans was cultured from 57% of the LJP family members accessioned into the study. Nine of 36 LJP-susceptible subjects, in seven families, developed signs of periodontal destruction. All but one of these conversion subjects harbored A. actinomycetemcomitans. Bacterial variants representative of a single RFLP group (II) showed the strongest correlation with conversion (P < 0.002). Six of nine conversion subjects were infected with A. actinomycetemcomitans from this group. RFLP group II variants also prevailed in 8 of 22 probands but were absent in the 35 healthy control subjects. In contrast to the selective distribution of group II variants is diseased individuals, variants belonging to RFLP groups XIII and XIV were found exclusively in the control subjects. Thus, the use of RFLP to type clinical isolates of A. actinomycetemcomitans has resulted in the identification of genetic variants that predominate in LJP and health. These results indicate that studies concerned with the pathogenicity of this bacterium in LJP should be focused on the group II variants.

Adult↗

PubMed1997

Differential recognition of members of the carcinoembryonic antigen family by Opa variants of Neisseria gonorrhoeae.

Opacity (Opa) protein variation in Neisseria gonorrhoeae is implicated in the pathogenesis of gonorrhea, possibly by mediating adherence and entry of the bacteria into human tissues. One particular Opa protein mediates adherence to epithelial cells through cell surface proteoglycans. Recently, two other eukaryotic cell receptors for Opa proteins have been reported. These receptors are members of a subgroup of the carcinoembryonic (CEA) gene family that express CD66 antigens. CEA family members vary in their distribution in human tissues. In order to understand whether interactions between Opa and CEA-like molecules play any role in pathogenesis, we must investigate which CEA family members are able to serve as Opa receptors and which Opa proteins recognize CEA-like molecules. We therefore studied HeLa cells that were stably transfected with five different members of the CEA family, i.e., CEA, CEA gene family member 1a (CGM1a), CGM6, nonspecific cross-reacting antigen (NCA), and biliary glycoprotein a (BGPa). We infected these transfectants with all possible 11 Opa variants of gonococcal strain MS11 and determined the numbers of bacteria that were bound and internalized. To account for proteoglycan-mediated adherence, infection assays were also performed in the presence of heparin. Our results show that of the 11 Opa variants of MS11, the same 4 recognized CGM1a and NCA. CGM6, however, was not recognized by any Opa variant of MS11. CEA was recognized by at least 9 of 11 Opa variants, and the BGP transfectants specifically bound and internalized 10 of 11 Opa variants and also bound Opa-negative gonococci. Immunofluorescence experiments showed that clustering of CEA-like molecules occurred upon infection of HeLa transfectants with those Opa variants that interacted specifically with the CEA family member. Together these data show that CEA family members are differentially recognized by gonococcal Opa variants, suggesting that this phenomenon may contribute to cell tropism displayed by gonococci.

Antigens, Bacterial↗