Mark D. Brennan

Evidence for two schizophrenia susceptibility genes on chromosome 22q13.

Objective Previous linkage scans and meta-analyses for schizophrenia susceptibility loci failed to include the most distal portion of chromosome 22q. Accordingly, 27 families having individuals affected with schizophrenia and schizophrenia-spectrum disorders were analyzed using a set of highly informative markers covering all of chromosome 22q. Methods Microsatellite and single nucleotide polymorphism markers were evaluated by nonparametric linkage, parametric linkage, and transmission disequilibrium testing of 22q. Results The maximum nonparametric logarithm of odd scores were 2.9 (P=0.0016) for schizophrenia and 2.7 (P=0.003) for a broader disease definition that included schizotypal personality disorder–both at 44.5 cM within the Sult4A1 locus. Parametric models assuming dominant modes of inheritance and genetic heterogeneity gave maximum multipoint logarithm of odd scores for the broader disease definition at the Sult4A1 locus of 3.3 (P=0.0006) and single point logarithm of odd scores of 3.1–4.8 for Sult4A1 markers (P=0.000015–0.0005). A distal locus, centered at 61 cM, shows a maximum nonparametric logarithm of odd scores of 1.5 (P=0.072) for the broader disease definition. Transmission disequilibrium testing for three adjacent microsatellite markers located near the distal linkage peak revealed significant values for marker D22s526 for schizophrenia (P=0.0016–0.14) and for broader disease definitions including schizotypal personality disorder (P=0.0002–0.0003), and both schizotypal personality disorder plus schizoaffective disorder (P=0.00001–0.000077). Conclusion At least two separable, but closely linked, loci within 22q13 influencing susceptibility to schizophrenia-spectrum disorders, might be possible.

High Throughput Genotyping Technologies for Pharmacogenomics

Genetic differences between individuals play a role in determining susceptibility to diseases as well as in drug response. The challenge today is first to discover the range of genetic variability in the human population and then to define the particular gene variants, or alleles, that contribute to clinically important outcomes. Consequently, high throughput, automated methods are being developed that allow rapid scoring of microsatellite alleles and single nucleotide polymorphisms (SNPs). Many detection technologies are being used to accomplish this goal, including electrophoresis, standard fluorescence, fluorescence polarization, fluorescence resonance energy transfer, and mass spectrometry. SNP alleles may be distinguished by any one of several methods, including single nucleotide primer extension, allele-specific hybridization, allele-specific primer extension, oligonucleotide ligation assay, and invasive signal amplification. Newer methods require less sample manipulation, increase sensitivity, allow more flexibility, and decrease reagent costs. Recent developments show promise for continuing these trends by combining amplification and detection steps and providing flexible, miniaturized platforms for genotyping.

Association of Sult4A1 SNPs with psychopathology and cognition in patients with schizophrenia or schizoaffective disorder

A number of genes located on chromosome 22q11-13, including catechol-O-methyltransferase (COMT), are potential schizophrenia susceptibility genes. Recently, the sulfotransferase-4A1 (Sult4A1) locus within chromosome 22q13 was reported to be linked to schizophrenia in a family TDT study. Sult4A1 is related to metabolism of monoamines, particularly dopamine and norepinephrine, both of which have been implicated in the pathophysiology of the psychopathology and cognitive dysfunction components of schizophrenia. An available, prospectively collected data base was interrogated to determine how three Sult4A1 SNPs: rs138060, rs138097, and rs138110, previously shown to be associated with schizophrenia might be associated with psychopathology, cognition, and quality of life in a sample of 86 Caucasian patients with schizophrenia or schizoaffective disorder. The majority of patients met criteria for treatment resistant schizophrenia and had been drug-free for one week or longer at the time of evaluation. The major findings were: 1) patients heterozygous (T/G) for rs138060 had significantly worse Brief Psychiatric Rating Scale (BPRS) Total and anxiety/depression sub-scale scores, and higher Scale for the Assessment of Positive Symptoms (SAPS) Total scores than G/G homozygous patients; and 2) patients heterozygous (A/G) for rs138097 demonstrated significantly worse performance on neuropsychological testing, specifically on tests of executive function and working memory, compared to patients homozygous for the G and A alleles. RS138110 was unrelated to psychopathology and cognition. These results provide the first evidence of how genetic variation in Sult4A1 may be related to clinical symptoms and cognitive function in schizophrenia, and permit future studies to attempt to replicate these potentially important findings.

The gypsy Insulator Can Act as a Promoter-Specific Transcriptional Stimulator

ABSTRACT Insulators define chromosomal domains such that an enhancer in one domain cannot activate a promoter in a different domain. We show that the Drosophila gypsy insulator behaves as acis-stimulatory element in the larval fat body. Transcriptional stimulation by the insulator is distance dependent, as expected for a promoter element as opposed to an enhancer. Stimulation of a test alcohol dehydrogenase promoter requires a binding site for a GATA transcription factor, suggesting that the insulator may be facilitating access of this DNA binding protein to the promoter. Short-range stimulation requires both the Suppressor of Hairy-wing protein and the Mod(mdg4)-62.7 protein encoded by the trithorax group gene mod(mdg4). In the absence of interaction with Mod(mdg4)-62.7, the insulator is converted into a short-range transcriptional repressor but retains some cis-stimulatory activity over longer distances. These results indicate that insulator and promoter sequences share important characteristics and are not entirely distinct. We propose that the gypsy insulator can function as a promoter element and may be analogous to promoter-proximal regulatory modules that integrate input from multiple distal enhancer sequences.

MUC1 and estrogen receptor α gene polymorphisms in dry eye patients

Dry eye syndrome is a collection of common disorders of multifactorial etiology. Although the epidemiology of dry eye has been well studied, reports of genetic patterns that might influence susceptibility to dry eye are few. We reported that the frequency of non-Sjögrens aqueous-deficient dry eye patients expressing only the MUC1/A splice variant of the mucin MUC1 may be lower than that of a normal control group [Imbert, Y., Darling, D.S., Jumblatt, M.M., Foulks, G.N., Couzin, E.G., Steele, P.S., Young, W.W., Jr., 2006. MUC1 splice variants in human ocular surface tissues: possible differences between dry eye patients and normal controls. Exp. Eye Res. 83, 493-501]. Also, He et al. [He, Y., Li, X., Bao, Y., Sun, J., Liu, J., 2006. The correlation of polymorphism of estrogen receptor gene to dry eye syndrome in postmenopausal women. Yan. Ke. Xue. Bao. 22, 233-236] reported a difference between Chinese dry eye and control groups in the frequency of a polymorphism in estrogen receptor alpha (ERalpha). In the present study we determined the statistical significance and generality of these observations and tested if the MUC1 splice variant difference between subject groups reflected a difference in the MUC1 variable number of tandem repeat (VNTR) size class. There was a perfect correlation between the MUC1/A or MUC1/B splice variant pattern and the SNP genotype frequency of the SNP (rs4072037) controlling that splicing event. In contrast, western and Southern blotting indicated that MUC1 VNTR size class corresponded to the MUC1 SNP genotype in only 80% of cases. We determined the status of the MUC1 SNP in normal and dry eye populations all of whom were female Caucasians. The MUC1 SNP genotype frequency of the normal control group was statistically different from both the non-Sjögrens aqueous-deficient dry eye group with ocular surface staining (P=0.017) and the evaporative dry eye group (P=0.015). We also tested SNP rs2234693 to analyze the polymorphism in the ERalpha gene and found no significant difference in the SNP genotype frequency between the control group and either of the dry eye subtypes. Thus, among Caucasians there is no evidence for an association of the ERalpha gene polymorphism with dry eye syndrome as previously described in a Chinese population. In conclusion, the etiologies of evaporative dry eye and non-Sjögrens aqueous-deficient dry eye are known to be different. However, our results suggest that both of these subtypes of dry eye disease may share a common mechanism or factor related to MUC1 genotypic differences that affects susceptibility to ocular surface damage. This altered susceptibility may not be related to the MUC1 VNTR size class. Therefore, mechanisms influencing protection of the ocular surface against inflammation and damage in different types of dry eye disease warrant further investigation particularly in relation to MUC1 genotype.

Evidence for a SULT4A1 haplotype correlating with baseline psychopathology and atypical antipsychotic response

AIM This study evaluated the impact of SULT4A1 gene variation on psychopathology and antipsychotic drug response in Caucasian subjects from the Clinical Antipsychotic Trials of Intervention Effectiveness (CATIE) study and a replication sample. PATIENTS & METHODS SULT4A1 haplotypes were determined using SNP data. The relationship to baseline psychopathology was evaluated using linear regression of Positive and Negative Syndrome Scale (PANSS) total score. Drug response was evaluated using Mixed Model Repeat Measures (MMRM) for change in PANSS. RESULTS For the CATIE sample, patients carrying a haplotype designated SULT4A1-1(+) displayed higher baseline PANSS (p = 0.03) and, when treated with olanzapine, demonstrated a significant interaction with time (p = 0.009) in the MMRM. SULT4A1-1(+) patients treated with olanzapine displayed improved response compared with SULT4A1-1(-) patients treated with olanzapine (p = 0.008) or to SULT4A1-1(+) patients treated with risperidone (p = 0.006). In the replication sample, SULT4A1-1(+) patients treated with olanzapine demonstrated greater improvement than SULT4A1-1(-) patients treated with olanzapine (p = 0.05) or than SULT4A1-1(+) patients treated with risperidone (p = 0.05). CONCLUSION If validated, determination of SULT4A1-1 haplotype status might be useful for identifying patients who show an enhanced response to long-term olanzapine treatment. Original submitted 6 October 2010; Revision submitted 9 December 2010.

Transmission disequilibrium suggests a role for the sulfotransferase-4A1 gene in schizophrenia

Previous studies suggest a role for chromosome 22q13 in schizophrenia. This segment of chromosome 22 contains the sulfotransferase‐4A1 (Sult4A1) gene, which encodes an enzyme thought to be involved in neurotransmitter metabolism in the central nervous system. To evaluate this candidate, we developed a microsatellite marker targeting a polymorphism in its 5′ nontranslated region (D22s1749E). Using samples obtained from the National Institutes of Mental Health Schizophrenia Genetics Initiative, we evaluated 27 families having multiple siblings with schizophrenia and schizophrenia‐spectrum disorders for transmission disequilibrium (TDT) of this marker along with three single nucleotide polymorphisms (SNPs) spanning a 37 kb segment containing the Sult4A1 gene. TDT for D22s1749E was significant (P < 0.05), with a tendency for the 213 nt allele to be preferentially transferred to affected children (P = 0.0079). Global chi‐square values for haplotypes involving the SNPs (ss146366, ss146407, and ss146420) and D22s1749E, also showed significant TDT values (P = 0.0006–0.0016). Consequently, we proposed that Sult4A merited more careful scrutiny as a candidate gene for schizophrenia susceptibility.

Complexity in evolved regulatory variation for alcohol dehydrogenase genes in HawaiianDrosophila

SummaryThe alcohol dehydrogenase gene (Adh gene) ofDrosophila affinidisjuncta is expressed at a higher level in the larval midgut and Malpighian tubules than the homologous gene fromDrosophila hawaiiensis. This study analyzed thecis-acting sequences responsible for these regulatory differences in larval tissues ofDrosophila melanogaster transformants. A series of 10 chimeric and deletedAdh genes was introduced into the germ line ofD. melanogaster, and tissue-specific expression levels were quantified by gel electrophoresis of tissue extracts. Sequences in the upstream region of the two genes had the strongest influence on enzyme production in the midgut and Malpighian tubules. Other sequence elements also showed effects, some of which were tissue specific. Most gene fragments displayed context-dependent effects, thus supporting the proposed model of polygenic regulation ofAdh gene expression.

Genotypic variation in the SV2C gene impacts response to atypical antipsychotics the CATIE Study

Pharmacogenetic (PGx) predictors of response would improve outcomes in antipsychotic treatment. Based on both biological rationale and prior evidence of an impact on Parkinsons disease, we conducted an association study for 106 SNPs in the synaptic vesicle protein 2C (SV2C) gene using genetic and treatment response data from the Clinical Trial of Antipsychotic Intervention Effectiveness (CATIE). We examined response to the atypical antipsychotics for Caucasian subjects in the blinded phases, Phases 1A, 1B, and 2, of CATIE with sample sizes as follows: olanzapine (N=134), quetiapine (N=124), risperidone (N=134), and ziprasidone (N=74). Response was defined as change in the Positive and Negative Syndrome Scale (PANSS) score using a mixed model repeat measures approach. Subjects homozygous for the T allele of rs11960832 displayed significantly worse response to olanzapine treatment, the only finding with study-wide significance (p=2.94×10(-5); false discovery rate=2.18×10(-2)). These subjects also displayed worse response to quetiapine with nominal significance (p=4.56×10(-2)). While no other SNP achieved study-wide significance, one SNP (rs10214163) influencing Parkinsons disease displayed nominally significant association with olanzapine and quetiapine response, while the second such SNP (rs30196) showed a statistical trend toward correlating with olanzapine and quetiapine response. Furthermore, both coding SNPs examined (rs31244 and rs2270927) displayed nominally significant correlations with treatment response: one for olanzapine (rs227092), and one for quetiapine (rs31244). The fact that multiple SNPs in SV2C may impact response to atypical antipsychotics suggests that further evaluation of SNPs in this gene as PGx predictors of antipsychotic response is warranted.

Similar tissue-specific expression of the Adh genes from different Drosophila species is mediated by distinct arrangements of cis-acting sequences

In Drosophila melanogaster transformants, the alcohol dehydrogenase (Adh) genes from D. affinidisjuncta and D. grimshawi show similar levels of expression except in the adult midgut where the D. affinidisjuncta gene is expressed about 10- to 20-fold more strongly. To study the arrangement of cis-acting sequences responsible for this regulatory difference, homologous restriction sites were used to create a series of chimeric genes that switched fragments from the 5′ and 3′ flanking regions of these two genes. Chimeric genes were introduced into the germ-line of D. melanogaster, and Adh gene expression was analyzed by measuring RNA levels. Various gene fragments in the promoter region and elsewhere influence expression in the adult midgut and in whole larvae and adults. Comparison of these results with earlier studies involving chimeras between the D. affinidisjuncta and D. hawaiiensis genes indicates that expression in the adult midgut is influenced by multiple regulatory sequences and that distinct arrangements of regulatory sequences can result in similar levels of expression both in the adult midgut and in the whole organism.