Marvin Schwalb

An oligonucleotide based array-CGH system for detection of genome wide copy number changes including subtelomeric regions for genetic evaluation of mental retardation†

Developmental delay (DD) and mental retardation (MR) are important child heath issues with a one percent prevalence. Karyotyping with or without subtelomeric FISH (fluorescent in situ hybridization), unless the phenotype of the patient suggests a specific aberration for a specific FISH assay, is the most common procedure in cytogenetic evaluation of MR/DD. In addition, there are several platforms utilizing microarray based comparative genomic hybridization technology (array‐CGH) for genetic testing. Array‐CGH can detect deletions or duplications in very small segments of chromosomes and the use of this technology is expected to increase the diagnostic yield. The major limitation of the current BAC based array technologies is the low resolution (∼1 Mb) of the chip and suboptimal coverage particularly in the subtelomeric regions. Our aim was to design a novel array‐CGH chip with high‐density of probes in the subtelomeric regions as well as to maintain sufficient density in other regions of the genome to provide comprehensive coverage for DD/MR. For this purpose, we used Human Genome CGH Microarray 44B chip (Agilent) as the template for the novel design. Using e‐array 4.0 (Agilent), one third of the probes were randomly removed from the array and replaced by 14,000 subtelomeric probes. The average density of the probe coverage is 125 kb and 250–400 probes interrogate subtelomeric regions. To evaluate the array, we tested 15 samples (including subtelomeric aberrations and other microdeletion syndromes), which were previously analyzed by karyotyping and/or FISH. The concordance rate between array results and previous results is 100%. In addition we detected two novel aberrations that were not detected by karyotyping. These results demonstrate the utility of this format of array‐CGH in detecting genome wide submicroscopic copy number changes as well as providing comprehensive coverage of all subteleomeric regions.

Comprehensive genome-wide comparison of DNA and RNA level scan using microarray technology for identification of candidate cancer-related genes in the HL-60 cell line

Genome-wide scans for DNA and RNA changes in the HL-60 cell line relative to normal leukocytes were conducted. Microarray-based comparative genome hybridization (CGH) studies were performed with the Spectral Genomics Human Bacterial Artificial Chromosome (BAC) 3MB system. Transcriptional measurements of approximately 12,500 human genes were monitored using Affymetrix U95A GeneChips. In HL-60, genomic DNA amplification of the 8q24 locus, trisomy 18, and deletions at loci 5q11.2 approximately q31, 6q12, 9p21.3 approximately p22, 10p12 approximately p15, 14q22 approximately q31, 17p12 approximately p13.3, and monosomy X were detected. After obtaining locus information about the RNA transcripts from the Affymetrix database, 4368 genes were stratified both according to status of RNA expression and the DNA copy number of their designated loci. The expression level of 2326 (53.25%) of 4368 transcripts is concordant with DNA copy number. Examples of specific, highly expressed, cancer-associated genes in amplified loci include SERPINB10, MYC, TYMS, HEC, and EPB41L3, while CD14, GZMK, TCF7, FOS, MLH3, CTNNA1, IRF1, VIM, CRK, MAP3K1, STAM, MAX, SFRG5, ENC1, PURA, MNT, RASA1, GLRX, UBE2B, NR3C1, PTENP1, BS69, COPEB, SKIP, PIM2, and MIC2 represent cancer-associated genes in deleted loci with decreased expression. The complementary usage of genome-wide DNA and RNA scans should enhance the identification of candidate genes in the neoplastic process.

Commitment to fruiting in synchronously developing cultures of the basidiomycete Schizophyllum commune

Summary1. A simple technique is described for obtaining large numbers of localized and synchronized fruit bodies of Schizophyllum commune by lowering the carbon dioxide level in mated cultures. 2. The fruiting process is unaffected by a continuous supply of glucose (0.4–2%). However, synchrony and commitment are lost when the glucose concentration is raised to 4%. 3. Five per cent carbon dioxide inhibits the further development of fruit bodies at all stages except the expansion of the already formed basidiocarp. 4. Fruit bodies can develop at 30°C when a CO2 trapping agent is employed, contrary to previous reports. 5. A possible relationship between the rate of vegetative growth and the initiation of fruiting is discussed.

The decision to test in women receiving genetic counseling for BRCA1 and BRCA2 mutations.

Functions of genetic counseling include provision of risk information and provision of support in an effort to assist with decision making. This study examines (1) the relationship among intentions to test, self-reported provision of blood sample, and receipt of test results; (2) the impact of genetic counseling on distress specific to gene status, perceived risk of developing breast and ovarian cancer in the context having BRCA1/2 mutations (mutations predisposing to increased risk of breast–ovarian cancer), and perceived risk factors for breast cancer; and (3) the clinical profile of those receiving/not receiving results. Intentions to test for BRCA1/2 mutations, self-reported provision of blood sample immediately after counseling, and receipt of test results were statistically different but highly correlated, and intentions to test increased from pre- to postcounseling. A repeated measures ANOVA found distress specific to gene status and perceived risk factors decreased from pre- to postcounseling. Further, two clinical profiles of consultands emerged: (1) those receiving results with change in intentions to test having lower levels of distress and (2) those not receiving results and those receiving results with no change in intentions to test with higher levels of distress. Our findings are consistent with the function of genetic counseling—to provide information and support to those with familial cancer, as well as to assist in decision making. The provision of support is important as distress specific to gene status may impede flexible decision making about genetic testing.

Effect of adenosine 3′,5′-cyclic monophosphate on the morphogenesis of fruit bodies ofSchizophyllum commune

Adenosine 3′,5′-cyclic monophosphate has been found to affect the normal development of fruit bodies in the basidiomyceteSchizophyllum commune. At a concentration of 10-3 M, cyclic AMP causes many fruit bodies to cease morphogenesis at an early stage. Those fruit bodies which continue development have either no gills or abnormal gills. A pronounced effect was also observed on the fruit bodies carrying a dominant mutant.

Cancer Genetics Knowledge and Beliefs and Receipt of Results in Ashkenazi Jewish Individuals Receiving Counseling for BRCA1/2 Mutations

BACKGROUND Genetic counseling for BRCA1 and BRCA2 mutations (mutations associated with increased risk of breast-ovarian cancer) endeavors to communicate information that will help individuals make informed decisions regarding genetic testing. METHODS This repeated-measures study examined cancer genetics knowledge and beliefs before and after counseling and their relationship to receipt of results for BRCA1/2 mutations in 120 highly educated Ashkenazi Jewish individuals. RESULTS A repeated-measures analysis examined change in knowledge and beliefs regarding personal behavior, mechanisms of cancer inheritance, meaning of a positive result, practitioner knowledge, frequency of inherited cancer, and meaning of a negative result from pre- to post-counseling with the between subjects variables of education (with/without graduate training) and personal history of breast or ovarian cancer (yes/no), and risk of having a mutation entered as a covariate. Mechanisms of cancer inheritance, meaning of a positive result, and practitioner knowledge increased from pre- to post-counseling. Those with graduate training had higher ratings of mechanisms of cancer inheritance ratings and lower ratings of frequency of inherited cancers than those without. Mann-Whitney U tests found those testing had higher ratings in mechanisms of cancer inheritance, specifically in the association of multiple primary cancers with hereditary cancer, than those not testing. CONCLUSIONS Genetic counseling is helpful in improving overall knowledge of cancer genetics even for highly educated individuals. Particular areas of knowledge improvement should be explored in relation to receipt of results, especially to further elucidate the relationship of knowledge of the association of multiple primary cancers with hereditary cancer to receipt of test results.

Changes in activity of enzymes metabolizing glucose 6-phosphate during development of the basidiomycete Schizophyllum commune

Abstract During the formation of fruit bodies in the basidiomycete Schizophyllum commune , there are changes in the specific activities of the enzymes utilizing glucose 6-phosphate. These include a 50% increase in the relative amount of glucose-6-phosphate dehydrogenase followed by a decrease in this enzyme and a 90% decrease in phosphoglucomutase activity. This results in a 10-fold change in the relationship between these two enzymes during development. The activities in several mutants were examined and abnormal enzyme levels were found in some cases. The activity of 6-phosphogluconate dehydrogenase was also examined, and its contribution to the assay of the other enzymes was determined.

Copy number variations in three children with sudden infant death

Sudden death of an infant is a devastating event that needs an explanation. When an explanation cannot be found, the case is labeled as sudden infant death syndrome or unclassified sudden infant death. The influence of genetic factors has been recognized for sudden infant death, but copy number variations (CNVs) as potential risk factors have not been evaluated yet. Twenty‐seven families were enrolled in this study. The tissue specimens from deceased children were obtained and array‐based comparative genomic hybridization (array‐CGH) experiments were performed on the genomic DNA isolated from these specimens using Agilent Technologies Custom 4 × 44K arrays. Quantitative polymerase chain reaction experiments were performed to confirm the overlapping duplication and deletion region in two different cases. A de novo CNV is detected in 3 of 27 cases (11%). In case 1, an ∼3‐Mb (chr 8: 143,211,215‐qter) duplication on 8q24.3‐qter and a 4.4‐Mb deletion on the 22q13.3‐qter (chr 22: 45,047,068‐qter) were detected. Subtelomeric chromosome analysis of the father and the surviving sibling of case 1 showed a balanced reciprocal translocation, 46,XY,t(8;22)(q24.3;q13.3). A 240‐kb (chr 6: 26,139,810‐26,380,787) duplication and a 1.9‐Mb deletion (chr 6: 26,085,971‐27,966,150) at chromosome 6p22 were found in cases 2 and 3, respectively. Array‐CGH and conventional cytogenetic studies did not reveal the observed CNVs in the parents and the siblings of cases 2 and 3. The detected CNVs in cases 2 and 3 encompassed several genes including the major histone cluster genes. Array‐CGH analysis may be beneficial during the investigations after sudden infant death.

Phenol oxidase activity during development of Coprinus cinereus

Coprinus cinereus produces a developmentally regulated phenol oxidase which appears to be responsible for the black pigmentation of basidiospores.

Variants of ST8SIA1 Are Associated with Risk of Developing Multiple Sclerosis

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system of unknown etiology with both genetic and environmental factors playing a role in susceptibility. To date, the HLA DR15/DQ6 haplotype within the major histocompatibility complex on chromosome 6p, is the strongest genetic risk factor associated with MS susceptibility. Additional alleles of IL7 and IL2 have been identified as risk factors for MS with small effect. Here we present two independent studies supporting an allelic association of MS with polymorphisms in the ST8SIA1 gene, located on chromosome 12p12 and encoding ST8 alpha-N-acetyl-neuraminide alpha-2,8-sialyltransferase 1. The initial association was made in a single three-generation family where a single-nucleotide polymorphism (SNP) rs4762896, was segregating together with HLA DR15/DQ6 in MS patients. A study of 274 family trios ( affected child and both unaffected parents) from Australia validated the association of ST8SIA1 in individuals with MS, showing transmission disequilibrium of the paternal alleles for three additional SNPs, namely rs704219, rs2041906, and rs1558793, with p = 0.001, p = 0.01 and p = 0.01 respectively. These findings implicate ST8SIA1 as a possible novel susceptibility gene for MS.