Biomedical subjects

Q Yang

Publications and source records attributed to Q Yang.

At least 217 records · Page 12Linked to original sources

PubMed1999

Inhibition of DNA methyltransferase stimulates the expression of signal transducer and activator of transcription 1, 2, and 3 genes in colon tumor cells.

Inhibitors of DNA methyltransferase, typified by 5-aza-2'-deoxycytidine (5-Aza-CdR), induce the expression of genes transcriptionally down-regulated by de novo methylation in tumor cells. We utilized gene expression microarrays to examine the effects of 5-Aza-CdR treatment in HT29 colon adenocarcinoma cells. This analysis revealed the induction of a set of genes that implicated IFN signaling in the HT29 cellular response to 5-Aza-CdR. Subsequent investigations revealed that the induction of this gene set correlates with the induction of signal transducer and activator of transcription (STAT) 1, 2, and 3 genes and their activation by endogenous IFN-alpha. These observations implicate the induction of the IFN-response pathway as a major cellular response to 5-Aza-CdR and suggests that the expression of STATs 1, 2, and 3 can be regulated by DNA methylation. Consistent with STAT's limiting cell responsiveness to IFN, we found that 5-Aza-CdR treatment sensitized HT29 cells to growth inhibition by exogenous IFN-alpha2a, indicating that 5-Aza-CdR should be investigated as a potentiator of IFN responsiveness in certain IFN-resistant tumors.

Antimetabolites, Antineoplastic↗

PubMed1999

Stabilization of liposomal membranes by thermozeaxanthins: carotenoid-glucoside esters.

Thermozeaxanthins (TZS) are novel carotenoid-glucoside esters existing in the cell membranes of the thermophilic bacterium, Thermus thermophilus. The effect of TZS on membrane permeability was studied by measuring the leakage of the fluorescent dye from calcein-entrapped large unilamellar liposomes (LUVs). The LUVs were composed of a small portion (0.2-1.0 mol%) of TZS and phosphatidylcholine (PC) of various length and saturation degree of hydrocarbon chains. Incorporation of TZS in egg PC LUVs stabilized the liposomes in the temperature range from 30 to 80 degrees C, as only 2.6% of the entrapped calcein leaked out in contrast to 10% release from the egg PC liposomes without TZS. LUVs composed of dipalmitoylphosphatidylcholine (DPPC) or dioleoylphosphatidylcholine (DOPC) were stabilized by the incorporation of TZS at a temperature below 30 degrees C. Inclusion of TZS in LUVs composed of dimyristoylphosphatidylcholine, whose hydrocarbon chains are shorter than both DPPC and DOPC, did not stabilize the liposomes. About 90% of the entrapped dye was lost indicating defects of the liposomal membranes. Matching of the lipid bilayer thickness with the molecular length of TZS in the bilayers is discussed.

Carotenoids↗

PubMed1999

In vivo modeling of myosin binding protein C familial hypertrophic cardiomyopathy.

Myosin binding protein C (MyBP-C) is an integral part of the striated muscle sarcomere. As is the case for other sarcomeric genes in human populations, multiple mutations within the gene have been linked to familial hypertrophic cardiomyopathy. Although some MyBP-C lesions are the result of missense mutations, most show truncated polypeptides lacking either the myosin or myosin and titin binding sites. Previously, we generated transgenic (TG) mice with cardiac-specific expression of a MyBP-C mutant lacking the myosin and titin binding domains. Surprisingly, the mutant protein was stable and made up a majority of the MyBP-C species, with concomitant reductions in endogenous MyBP-C such that overall MyBP-C stoichiometry was conserved. In the present study, we created a second series of TG mice that express, in the heart, a mutant MyBP-C lacking only the myosin binding site. In contrast to the previous data for the MyBP-C lacking both titin and myosin binding sites, only very modest levels of protein were found, consistent with data obtained from human biopsies in which mutated MyBP-C could not be detected. Despite normal levels of wild-type MyBP-C, there were significant changes in the structure and ultrastructure of the heart. Fiber mechanics showed decreased unloading shortening velocity, maximum shortening velocity, and relative maximal power output.

Animals↗

PubMed1999

Domain structure of gpNu1, a phage lambda DNA packaging protein.

The terminase enzyme from bacteriophage lambda is responsible for the insertion of a dsDNA genome into the confines of the viral capsid. The holoenzyme is composed of gpA and gpNu1 subunits in a gpA(1) x gpNu1(2) stoichiometry. While genetic studies have described regions within the two proteins responsible for DNA binding, capsid binding, and subunit interactions in the holoenzyme complex, biochemical characterization of these domains is limited. We have previously described the cloning, expression, and biochemical characterization of a soluble DNA binding domain of the terminase gpNu1 subunit (Met1 to Lys100) and suggested that the hydrophobic region spanning Lys100 to Pro141 defines a domain responsible for self-association interactions, and that is important for cooperative DNA binding [Yang et al. (1999) Biochemistry 38, 465-477]. We further suggested that the genetically defined gpA-interactive domain in the C-terminal half of the protein is limited to the C-terminal approximately 40 amino acids of gpNu1. Here we describe the cloning, expression, and biochemical characterization of gpNu1DeltaP141, a deletion mutant of gpNu1 that comprises the DNA binding domain and the putative hydrophobic self-assembly domain of the full-length protein. Purified gpNu1DeltaP141 shows a strong tendency to aggregate in solution; However, the protein remains soluble in 0.4 M guanidine hydrochloride, and circular dichroism (CD) and fluorescence spectroscopic studies demonstrate that the protein is folded under these conditions. Moreover, CD spectroscopy and thermally induced unfolding studies suggest that the DNA binding domain and the self-association domain represent independent folding domains of gpNu1DeltaP141. The mutant protein interacts weakly with the gpA subunit, but does not form a catalytically competent holoenzyme complex, suggesting that the C-terminal 40 residues are important for appropriate subunit interactions. Importantly, gpNu1DeltaP141 binds DNA tightly, but with less specificity than does full-length protein, and the data suggest that the C-terminal residues are further required for specific DNA binding activity. The implications of these results in the assembly of a functional holoenzyme complex are discussed.

Bacteriophage lambda↗

PubMed1999

Tubulointerstitial nephritis antigen: an extracellular matrix protein that selectively regulates tubulogenesis vs. glomerulogenesis during mammalian renal development.

Tubulointerstitial nephritis antigen (TIN-ag) is an extracellular matrix protein and is expressed in the renal tubular basement membranes. Its role in metanephric development was investigated. TIN-ag cDNA, isolated from the newborn mouse library, had an ORF of 1,425 nucleotides, a putative signal sequence, and an ATP/GTP-binding site. The translated sequence had approximately 80% identity with rabbit TIN-ag. Among various tissues, TIN-ag mRNA was primarily expressed in the newborn kidney. In the embryonic metanephros, TIN-ag expression was confined to the distal convolution or pole of the S-shaped body, the segment of the nascent nephron that is the progenitor of renal tubules. Treatment with TIN-ag antisense oligodeoxynucleotide induced dysmorphogenesis of the embryonic metanephroi, malformation of the S-shaped body, and a decrease in the tubular population, whereas the glomeruli were unaffected. Treatment also led to a decrease of TIN-Ag mRNA, de novo synthesis of TIN-ag protein, and its antibody reactivity. The mRNA expression of glomerular epithelial protein 1 (a marker for renal podocytes), anti-heparan-sulfate-proteoglycan antibody reactivity, and wheat germ agglutinin lectin staining of the metanephros were unaffected. The anti-TIN-ag antibody treatment also caused deformation of the S-shaped body and a reduction in the tubular population, whereas the glomeruli were unchanged. The data suggest that the TIN-ag, unlike other basement membrane proteins, selectively regulates tubulogenesis, whereas glomerulogenesis is largely unaffected.

Amino Acid Sequence↗

PubMed1999

Human insulin gene is a target gene of hepatocyte nuclear factor-1alpha (HNF-1alpha) and HNF-1beta.

Maturity-onset diabetes of the young (MODY) is a monogenic form of diabetes characterized by autosomal dominant inheritance, early-onset, and impaired insulin secretion. The type 3 and type 5 forms of MODY result from mutations in the genes encoding the transcription factor, hepatocyte nuclear factor (HNF)-1alpha and HNF-1beta, respectively. The mechanism by which mutations in one allele of the HNF-1 gene impair pancreatic beta cell function is unclear. We studied the effects of wild-type and four mutant (L12H, R263C, P379fsdelCT, and L584S585fsinsTC) HNF-1alpha, which were identified in Japanese subjects with MODY3 on human insulin gene transcription. Both wild-type (WT) HNF-1alpha and HNF-1beta bound to the oligonucleotide containing the A3 element sequence in the human insulin promoter and transactivated the insulin-luciferase reporter gene by 30- and 31-fold, respectively. In contrast, binding of L12H, R263C and L584S585fsinsTC-HNF-1alpha to the probe was impaired. Transactivation activity by the four mutant HNF-1alpha was reduced (4.3 to 43.3% of WT). These data suggest that the insulin gene is a candidate target gene of HNF-1alpha/HNF-1beta and the impairment of insulin gene transcription by mutations in the HNF-1 gene might be involved in the pathogenesis of MODY.

Animals↗

PubMed1999

Cloning of rat fibrillin-2 cDNA and its role in branching morphogenesis of embryonic lung.

Fibrillin-2 is an extracellular matrix protein. It is associated with elastic fibers in several tissues and is believed to serve as a ligand for alphavbeta3 integrin, the latter being a known morphogen. In this study, the role of fibrillin-2 in lung development was investigated. Also, rat fibrillin-2 cDNA was isolated and sequenced and its spatiotemporal expression determined. It had approximately 88% homology with human fibrillin-2 and had Ca(2+) binding epidermal growth factor-like domains, transforming growth factor-beta binding protein motifs, and two RGD binding sites. Northern blot analysis revealed an approximately 10-kb transcript, and fibrillin-2 expression was developmentally regulated, and it paralleled that of tropoelastin. At day 13 of gestation, fibrillin-2 was expressed in the mesenchyme and at the epithelial:mesenchymal interface. From day 13 to 19 of gestation, its expression intensified and was confined around the tracheobronchial airways, while it lessened during the postnatal period. Immunoprecipitation revealed an approximately 350-kDa band by SDS-PAGE. Treatment with fibrillin-2 antisense oligodeoxynucleotide induced dysmorphogenesis of the lung explants. They were smaller and had rudimentary lung bud branches, collapsed conducting airways, and loose expanded mesenchyme. Concomitantly, fibrillin-2 mRNA, antibody reactivity in the explants, and fibrillin-2-specific radioincorporation were reduced. Anti-alphav and -laminin antibody reactivity and their respective incorporated specific radioactivities were unaltered. These data indicate that fibrillin-2 modulates organogenesis of the lung in the context of epithelial:mesenchymal interactions. Conceivably, the collapse of the conducting airways may also be related to the perturbed biology of the fibrillin-2 interacting protein, i.e., elastin, the latter being critical for the normal biophysiology of the lungs.

Actins↗

PubMed1999

Intraductal spread of invasive breast carcinoma has a positive correlation with c-erb B-2 overexpression and vascular invasion.

BACKGROUND: Studies of the histologic characteristics and biologic behavior of the intraductal spread of breast carcinoma are critically important in that they may lead to the identification of a unique spread pattern rather than a noninvasive lesion. METHODS: Paraffin embedded specimens of 187 primary invasive breast carcinomas and 4 noninvasive ductal carcinomas, obtained by wide excision, quadrantectomy, total glandectomy, or mastectomy, were studied immunohistochemically. The overexpression of c-erb B-2, p53, bcl-2, and MIB-1, as well as the histologic characteristics of intraductal spread (such as histologic features and histologic grade), were assessed. Chi-square and Fisher exact tests were conducted to evaluate significant differences; the Macintosh for Expert StatView 4.0 system was used to conduct these tests. RESULTS: The histologic characteristics of intraductal spread were similar to those of noninvasive ductal carcinoma. However, the expressions of c-erb B-2, p53, and other biologic markers of intraductal spread were similar to those of the main invasive tumor. The overexpression of c-erb B-2 protein was found more often in the group that was positive for intraductal spread than in the group that was negative (P < 0.01). Intraductal spread was found more often in the group that was positive for lymphatic and venous invasion than in the group that was negative (P < 0.005). Subnipple margin positive status was related closely to intraductal spread (P < 0.0001). CONCLUSIONS: The positive correlation between intraductal spread and c-erb B-2 overexpression as well as lymphatic, venous invasion was recognized, and it was determined that intraductal spread of invasive breast carcinoma possesses an invasive and metastatic potential that is distinct from noninvasive ductal carcinoma.

Antigens, Nuclear↗

PubMed1999

Evaluation of Axillary Lymph Node Status in Breast Cancer with MRI.

BACKGROUND: We performed a retrospective study to establish the optimal radiological criteria for axillary lymph node metastases from breast cancer by measuring all dissected nodes, and to determine whether magnetic resonance imaging (MRI) could reliably reveal axillary involvement. METHODS: Pathological findings and MRI scans of 202 patients with invasive breast cancer were reviewed. The long- and short-axis dimensions of all level I an II lymph nodes were measured microscopically, and then the long-to-short axis (L/S) ratio of each node was calculated. These parameters were compared with pathological nodal status to define radiological criteria for axillary involvement. MRI was carried out using T1-weighted spin-eho sequences in the coronal and sagittal planes. On MRI, every detected lymph node was measured and the shape ofthe nodal cortex was also examined. Then the diagnostic ability of MRI was assessed using these morphologic criteria. RESULTS: On histopathological examinations of 4043 dissected lymph nodes, a long-axis dimension of 10 mm or larger combined with a long-to-short axis ratio of less than 1.6 was the most accurate criteria for predicting lymph node metastases. On MRI, eccentric cortical hypertrophy was seen in only metastatic axillae.When these morphologic features were used as criteria for malignancy, MRI had asensi-tivity of 79%, a specificity of 93%, and an accuracy of 88%. In 16 of 17 false-negative axillae, MRI showed normally sized lymph nodes (< 10 mm). CONCLUSION: Our study indicates that MRI is a useful diagnostic method for the evaluation of axillary nodal status, but is limited in the detection of small metastatic lymph nodes.

Journal Article↗

PubMed1999

Conformational epitope of the type III group B Streptococcus capsular polysaccharide.

The protective epitope of the type III group B streptococcal polysaccharide (GBSPIII) is length dependent and conformational. To obtain a more accurate characterization of the conformational epitope, ELISA inhibition and surface plasmon resonance studies were conducted on two GBSPIII-specific mAbs using a large panel of oligosaccharide probes. The results of the studies confirmed that 2 repeating units (RU) is the minimum binding unit and that, while increases in chain length from 2 RU to 7 RU caused further optimization of the epitope, it remained monovalent. A 3-fold increase in affinity was observed between 7 RU and 20 RU, which, by surface plasmon resonance studies on a Fab, was shown to be due to both further optimization of the individual epitope and the occurrence of multivalency of epitope. The data support our hypothesis that the conformational epitope is an extended helical segment of the GBSPIII. GBSPIII exists mainly in the random coil form, which structurally mimics short oligosaccharide self Ags, but it can infrequently and spontaneously form extended helices. Although not prevalent in GBSPIII, the immune system preferentially selects these helical epitopes because they are unique to the polysaccharide. Contrary to a previously proposed model of GBSPIII binding in which the binding of the first Ab propagates a continuum of helical epitopes, our binding kinetics are consistent only with the helical epitope's being discontinuous and infrequent.

Animals↗

PubMed1999

Transmission disequilibrium test (TDT) when only one parent is available: the 1-TDT.

The transmission disequilibrium test (TDT) is a useful method to locate mutations linked to disease genes associated with complex diseases. TDT requires genotypes of affected individuals and their parents. Recently, Ewens and Spielman (Am J Hum Genet 1998;62:450-8) extended the TDT for use in sibships with at least one affected and one unaffected individual and devised a new test called the sib transmission/disequilibrium test (S-TDT). The S-TDT can be applied to diseases with late age at onset such as non-insulin-dependent diabetes mellitus, psychiatric disorders, and diseases related to aging. For some disorders, it might be relatively easy to obtain the genotype of one parent either because the other parent is not available for study or he/she is not cooperative. Curtis and Sham (Ann Hum Genet 1995;59:323-36) showed that bias in transmitting certain alleles is introduced if only heterozygous parents and homozygous offspring are used in the TDT. In this paper, the authors propose a new test, the 1-TDT, to detect linkage between a candidate locus and a disease locus using genotypes of affected individuals and only one available parent for each affected individual. The test is not biased under the null hypothesis of no linkage or association. The authors validate their test using both simulated and real data sets. Finally, they show how to combine data from different types of families.

Age of Onset↗

PubMed1999

Absent middle cerebral artery flow predicts the presence and evolution of the ischemic penumbra.

OBJECTIVES: In acute ischemic stroke the pattern of a perfusion-imaging (PI) lesion larger than the diffusion-weighted imaging (DWI) lesion may be a marker of the ischemic penumbra. We hypothesized that acute middle cerebral artery (MCA) occlusion would predict the presence of presumed "penumbral" patterns (PI > DWI), ischemic core evolution, and stroke outcome. METHODS: Echoplanar PI, DWI, and magnetic resonance angiography (MRA) were performed in 26 patients with MCA territory stroke. Imaging and clinical studies (Canadian Neurological Scale, Barthel Index, and Rankin Scale) were performed within 24 hours of onset and repeated at days 4 and 90. RESULTS: MCA flow was absent in 9 of 26 patients. This was associated with larger acute PI and DWI lesions, greater PI/DWI mismatch, early DWI lesion expansion, larger final infarct size, worse clinical outcome (p < 0.01) and provided independent prognostic information (multiple linear regression analysis, p < 0.05). Acute penumbral patterns were present in 14 of 26 patients. Most of these patients (9 of 14) had no MCA flow, whereas all nonpenumbral patients (PI < or = DWI lesion) had MCA flow (p < 0.001). Penumbral-pattern patients with absent MCA flow had greater DWI lesion expansion (p < 0.05) and worse clinical outcome (Rankin Scale score, p < 0.05). CONCLUSIONS: Absent MCA flow on MRA predicts the presence of a presumed penumbral pattern on acute PI and DWI and worse stroke outcome. Combined MRA, PI, and DWI can identify individual patients at risk of ischemic core progression and the potential to respond to thrombolytic therapy beyond 3 hours.

Adult↗

PubMed1999

[Detection of ABO genotypes by simultaneous PCR-RFLP method]

OBJECTIVE: To study human ABO genotyping by means of multiplex PCR and restriction fragment length polymorphism. METHODS: Two specific fragments of ABO gene were simultaneously amplified in a single tube, and then the double restriction digestion with RE Kpn I and Alu I was performed in the same tube. The amplified products were analyzed by PAGE and silver staining. A total of 125 Han unrelated individuals living in Wuhan were genotyped. RESULTS: Six genotypes of ABO were detected and the distribution was in good agreement with Hardy-Weinberg equilibrium. CONCLUSION: The results demonstrate that PCR-RFLP based approach is convenient, reliable and should be of value in forensic application.

Journal Article↗

PubMed1999

Enhancement of UV-induced cytotoxicity by the adeno-associated virus replication proteins.

Adeno-associated virus (AAV) normally requires co-infection of a helper virus to complete its life cycle. However, under conditions of cellular stress, such as treatment with carcinogens or ultraviolet (UV) light, a permissive intracellular environment is established and AAV completes its replicative cycle producing low levels of progeny virus. AAV DNA replication is dependent upon viral replication proteins, Rep78 and Rep68. The detailed mechanism by which these proteins interact with host cell factors is unknown. We have used a cell line (Neo6) that inducibly expresses the AAV Rep proteins to study their effects on cells that have undergone UV-induced DNA damage. Induction of Rep protein expression immediately after a sub-lethal dose of UV irradiation resulted in rapid cell killing. Those cells that die had chromatin condensation while cellular membranes remained intact, suggesting that concurrent Rep expression and UV damage induces an apoptosis-like response. However, we did not observe any DNA degradation. Thus we believe that the combination of Rep expression and UV irradiation induces cell death that shares some of the characteristics of apoptosis. UV irradiation and Rep expression induced an increase in the level of the CDK inhibitor, p21Cip, and the appearance of modified forms of both p21Cip and Bcl-2. Alteration of normal expression of these cytostatic/apoptotic proteins provides insight into the intracellular targets of the AAV replication proteins.

Apoptosis↗

PubMed1999

Covalent immobilization of unilamellar liposomes in gel beads for chromatography.

For immobilized (proteo)liposome chromatography, unilamellar liposomes were covalently bound within gel beads that had been activated by CNBr, N-hydroxysuccinimide, tresyl, or chloroformate. Liposomes composed of phosphatidylcholine (PC) and 2 mol% of amino-containing lipid (phosphatidylethanolamine-caproylamine) were immobilized in the activated gels at 5-35 micromol lipid/ml gel and yields of 11-70%. The highest immobilized amount was found in chloroformate-activated TSK G6000PW gel, which contains large pore size (>100 nm). Liposomes composed of PC alone could also be attached to the chloroformate-activated gels at 33-42 micromol/ml gel and yields of 58-65%, probably by crosslinking of the phosphate moiety of phospholipid with the active group of the adsorbent. Liposomes prepared by various phospholipids with or without amino-containing lipids can generally be immobilized in the chloroformate-activated gels. The covalently bound liposomes were characterized by their high stability, unilamellarity, permeability of the membranes, and drug-membrane partition properties. A stable membrane phase was constructed for chromatographic experiments to be performed under extreme elution conditions.

Chromatography, Gel↗

PubMed1999

Chemokine and chemokine receptor gene variants and risk of non-Hodgkin's lymphoma in human immunodeficiency virus-1-infected individuals.

Normal B-lymphocyte maturation and proliferation are regulated by chemotactic cytokines (chemokines), and genetic polymorphisms in chemokines and chemokine receptors modify progression of human immunodeficiency virus-1 (HIV-1) infection. Therefore, 746 HIV-1-infected persons were examined for associations of previously described stromal cell-derived factor 1 (SDF-1) chemokine and CCR5 and CCR2 chemokine receptor gene variants with the risk of B-cell non-Hodgkin's lymphoma (NHL). The SDF1-3'A chemokine variant, which is carried by 37% of whites and 11% of blacks, was associated with approximate doubling of the NHL risk in heterozygotes and roughly a fourfold increase in homozygotes. After a median follow-up of 11.7 years, NHL developed in 6 (19%) of 30 SDF1-3'A/3'A homozygotes and 22 (10%) of 202 SDF1-+/3'A heterozygotes, compared with 24 (5%) of 514 wild-type subjects. The acquired immunodeficiency syndrome (AIDS)-protective chemokine receptor variant CCR5-triangle up32 was highly protective against NHL, whereas the AIDS-protective variant CCR2-64I had no significant effect. Racial differences in SDF1-3'A frequency may contribute to the lower risk of HIV-1-associated NHL in blacks compared with whites. SDF-1 genotyping of HIV-1-infected patients may identify subgroups warranting enhanced monitoring and targeted interventions to reduce the risk of NHL.

Acquired Immunodeficiency Syndrome↗

PubMed1999

Partitioning of triphenylalkylphosphonium homologues in gel bead-immobilized liposomes: chromatographic measurement of their membrane partition coefficients.

Unilamellar liposomes of small or large size, SUVs and LUVs, respectively, were stably immobilized in the highly hydrophilic Sepharose 4B or Sephacryl S-1000 gel beads as a membrane stationary phase for immobilized liposome chromatography (ILC). Lipophilic cations of triphenylmethylphosphonium and tetraphenylphosphonium (TPP+) have been used as probes of the membrane potential of cells. Interaction of TPP+ and triphenylalkylphosphonium homologues with the immobilized liposomal membranes was shown by their elution profiles on both zonal and frontal ILC. Retardation of the lipophilic cations on the liposome gel bed was increased as the hydrophobicity of the cations increased, indicating the partitioning of lipophilic cations into the hydrocarbon region of the membranes. The cations did not retard on the Sepharose or Sephacryl gel bed without liposomes, confirming that the cations only interact with the immobilized liposomes. Effects of the solute concentration, flow rate, and gel-matrix substance on the ILC were studied. The stationary phase volume of the liposomal membranes was calculated from the volume of a phospholipid molecule and the amount of the immobilized phospholipid, which allowed us to determine the membrane partition coefficient (KLM) for the lipophilic cations distributed between the aqueous mobile and membrane stationary phases. The values of KLM were generally increased with the hydrophobicity of the solutes increased, and were higher for the SUVs than for the LUVs. The ILC method described here can be applied to measure membrane partition coefficients for other lipophilic solutes (e.g., drugs).

Cations↗

PubMed1999

Demonstration of heterodimer formation between S100B and S100A6 in the yeast two-hybrid system and human melanoma.

S100B (S100beta) and S100A6 (calcyclin) are two 10-kDa Ca2+- and Zn2+-binding proteins coexpressed in melanoma and cell-cycle regulated. These proteins are members of the S100 subfamily and are thought to exert their function through interaction with intracellular target proteins. In order to search for potential target proteins interacting with S100B, we used a yeast two-hybrid strategy with human S100B as bait to screen a human brain cDNA library. The fusion proteins interacting with the S100B bait were identified as S100B, S100A1, and S100A6. This indicates the potential of S100B to form homodimers and heterodimers with other members of the S100 subfamily. By Northern and Western blotting, S100B and S100A6 were shown to be expressed at high levels in a panel of human melanoma cell lines. S100B and S100A6 were coimmunoprecipitated from melanoma cell lysates in the presence of 100 microM Zn2+. Confocal microscopy demonstrated that both proteins were distributed throughout the cytoplasm and concentrated in the nucleus. The demonstration of an association and colocalization of S100B and S100A6 in melanoma supports the possibility that an S100B/S100A6 heterodimer plays a functional role in these cells.

Animals↗