A systematic review and meta-analysis of germline BRCA mutations in pancreatic cancer patients identifies global and racial disparities in access to genetic testing.

BACKGROUND: Germline BRCA1 and BRCA2 mutations (gBRCAm) can inform pancreatic cancer (PC) risk and treatment but most of the available information is derived from white patients. The ethnic and geographic variability of gBRCAm prevalence and of germline BRCA (gBRCA) testing uptake in PC globally is largely unknown. MATERIALS AND METHODS: We carried out a systematic review and prevalence meta-analysis of gBRCA testing and gBRCAm prevalence in PC patients stratified by ethnicity. The main outcome was the distribution of gBRCA testing uptake across diverse populations worldwide. Secondary outcomes included: geographic distribution of gBRCA testing uptake, temporal analysis of gBRCA testing uptake in ethnic groups, and pooled proportion of gBRCAm stratified by ethnicity. The study is listed under PROSPERO registration number #CRD42022311769. RESULTS: A total of 51 studies with 16 621 patients were included. Twelve of the studies (23.5%) enrolled white patients only, 10 Asians only (19.6%), and 29 (56.9%) included mixed populations. The pooled prevalence of white, Asian, African American, and Hispanic patients tested per study was 88.7%, 34.8%, 3.6%, and 5.2%, respectively. The majority of included studies were from high-income countries (HICs) (64; 91.2%). Temporal analysis showed a significant increase only in white and Asians patients tested from 2000 to present (P < 0.001). The pooled prevalence of gBRCAm was: 3.3% in white, 1.7% in Asian, and negligible (<0.3%) in African American and Hispanic patients. CONCLUSIONS: Data on gBRCA testing and gBRCAm in PC derive mostly from white patients and from HICs. This limits the interpretation of gBRCAm for treating PC across diverse populations and implies substantial global and racial disparities in access to BRCA testing in PC.

Large scale calcium channel gene rearrangements in episodic ataxia and hemiplegic migraine: implications for diagnostic testing.

BACKGROUND: Episodic ataxia type 2 (EA2) and familial hemiplegic migraine type 1 (FHM1) are autosomal dominant disorders characterised by paroxysmal ataxia and migraine, respectively. Point mutations in CACNA1A, which encodes the neuronal P/Q-type calcium channel, have been detected in many cases of EA2 and FHM1. The genetic basis of typical cases without CACNA1A point mutations is not fully known. Standard DNA sequencing methods may miss large scale genetic rearrangements such as deletions and duplications. The authors investigated whether large scale genetic rearrangements in CACNA1A can cause EA2 and FHM1. METHODS: The authors used multiplex ligation dependent probe amplification (MLPA) to screen for intragenic CACNA1A rearrangements. RESULTS: The authors identified five previously unreported large scale deletions in CACNA1A in seven families with episodic ataxia and in one case with hemiplegic migraine. One of the deletions (exon 6 of CACNA1A) segregated with episodic ataxia in a four generation family with eight affected individuals previously mapped to 19p13. In addition, the authors identified the first pathogenic duplication in CACNA1A in an index case with isolated episodic diplopia without ataxia and in a first degree relative with episodic ataxia. CONCLUSIONS: Large scale deletions and duplications can cause CACNA1A associated channelopathies. Direct DNA sequencing alone is not sufficient as a diagnostic screening test.

Autosomal-dominant GTPCH1-deficient DRD: clinical characteristics and long-term outcome of 34 patients.

BACKGROUND: An autosomal dominantly inherited defect in the GCH1 gene that encodes guanosine triphosphate cyclohydrolase 1 (GTPCH1) is the most common cause of dopa-responsive dystonia (DRD). A classic phenotype of young-onset lower-limb dystonia, diurnal fluctuations and excellent response to levodopa has been well recognised in association with GCH1 mutations, and rare atypical presentations have been reported. However, a number of clinical issues remain unresolved including phenotypic variability, long-term response to levodopa and associated non-motor symptoms, and there are limited data on long-term follow-up of genetically proven cases. METHODS: A detailed clinical evaluation of 34 patients (19 women, 15 men), with confirmed mutations in the GCH1 gene, is presented. RESULTS AND CONCLUSIONS: The classic phenotype was most frequent (n = 23), with female predominance (F:M = 16:7), and early onset (mean 4.5 years) with involvement of legs. However, a surprisingly large number of patients developed craniocervical dystonia, with spasmodic dysphonia being the predominant symptom in two subjects. A subset of patients, mainly men, presented with either a young-onset (mean 6.8 years) mild DRD variant not requiring treatment (n = 4), or with an adult-onset (mean 37 years) Parkinson disease-like phenotype (n = 4). Two siblings were severely affected with early hypotonia and delay in motor development, associated with compound heterozygous GCH1 gene mutations. The study also describes a number of supplementary features including restless-legs-like symptoms, influence of female sex hormones, predominance of tremor or parkinsonism in adult-onset cases, initial reverse reaction to levodopa, recurrent episodes of depressive disorder and specific levodopa-resistant symptoms (writer's cramp, dysphonia, truncal dystonia). Levodopa was used effectively and safely in 20 pregnancies, and did not cause any fetal abnormalities.

Pollen grains in lake sediments: redeposition caused by seasonal water circulation.

Annual pollen deposition per unit area measured in sediment traps is two to four times greater than deposition measured in surface sediment cores. The difference is due to repeated redeposition of pollen from the sediment surface during seasons of water circulation. This process reduces variations in the percentages of different pollen types in sediment, but causes differences in the total amount of pollen accumulated in various parts of the lake basin.

Range shifts and adaptive responses to Quaternary climate change.

Tree taxa shifted latitude or elevation range in response to changes in Quaternary climate. Because many modern trees display adaptive differentiation in relation to latitude or elevation, it is likely that ancient trees were also so differentiated, with environmental sensitivities of populations throughout the range evolving in conjunction with migrations. Rapid climate changes challenge this process by imposing stronger selection and by distancing populations from environments to which they are adapted. The unprecedented rates of climate changes anticipated to occur in the future, coupled with land use changes that impede gene flow, can be expected to disrupt the interplay of adaptation and migration, likely affecting productivity and threatening the persistence of many species.

GABA and glutamate depolarize cortical progenitor cells and inhibit DNA synthesis.

We have found that, during the early stages of cortical neurogenesis, both GABA and glutamate depolarize cells in the ventricular zone of rat embryonic neocortex. In the ventricular zone, glutamate acts on AMPA/kainate receptors, while GABA acts on GABAA receptors. GABA induces an inward current at resting membrane potentials, presumably owing to a high intracellular Cl- concentration maintained by furosemide-sensitive Cl- transport. GABA and glutamate also produce increases in intracellular Ca2+ in ventricular zone cells, in part through activation of voltage-gated Ca2+ channels. Furthermore, GABA and glutamate decrease the number of embryonic cortical cells synthesizing DNA. Depolarization with K+ similarly decreases DNA synthesis, suggesting that the neurotransmitters act via membrane depolarization. Applied alone, GABAA and AMPA/kainate receptor antagonists increase DNA synthesis, indicating that endogenously released amino acids influence neocortical progenitors in the cell cycle. These results demonstrate a novel role for amino acid neurotransmitters in regulating neocortical neurogenesis.

Multiple mitochondrial DNA deletions in monozygotic twins with OPMD.

BACKGROUND: Oculopharyngeal muscular dystrophy (OPMD) is caused by expansions of the poly (A) binding protein 2 (PABP2) gene. Previous histological analyses have revealed mitochondrial abnormalities in the muscles of OPMD patients but their significance remains uncertain. OBJECTIVE: We had the rare opportunity to study monozygotic twins with identical expansions of the PABP2 gene but with markedly different severities of OPMD. Both had histological features of mitochondrial myopathy. We determined whether mitochondrial DNA abnormalities underlay these changes. METHODS: Clinical information was obtained by history and examination. Muscle biopsies were obtained from each subject and genetic analysis was performed using long-range PCR and Southern blotting. RESULTS: We demonstrate, for the first time, the presence of mitochondrial DNA (mtDNA) deletions by Southern blotting in individuals with OPMD. This correlates with the presence of mitochondrial myopathy in both twins. Moreover, both twins had different mtDNA deletions, which might explain their phenotypic differences. CONCLUSION: We hypothesise that mitochondrial dysfunction may occur as a consequence of PABP2 gene mutations, and that this dysfunction may affect the phenotypic manifestations of OPMD.

Chloride channel myotonia: exon 8 hot-spot for dominant-negative interactions.

Myotonia congenita (MC) is the commonest genetic skeletal muscle ion channelopathy. It is caused by mutations in CLCN1 on chromosome 7q35, which alter the function of the major skeletal muscle voltage-gated chloride channel. Dominant and recessive forms of the disease exist. We have undertaken a clinical, genetic and molecular expression study based upon a large cohort of over 300 UK patients. In an initial cohort of 22 families, we sequenced the DNA of the entire coding region of CLCN1 and identified 11 novel and 11 known mutations allowing us to undertake a detailed genotype-phenotype correlation study. Generalized muscle hypertrophy, transient weakness and depressed tendon reflexes occurred more frequently in recessive than dominant MC. Mild cold exacerbation and significant muscle pain were equally common features in dominant and recessive cases. Dominant MC occurred in eight families. We noted that four newly identified dominant mutations clustered in exon 8, which codes for a highly conserved region of predicted interaction between the CLC-1 monomers. Expressed in Xenopus oocytes these mutations showed clear evidence of a dominant-negative effect. Based upon the analysis of mutations in this initial cohort as well as a review of published CLCN1 mutations, we devised an exon hierarchy analysis strategy for genetic screening. We applied this strategy to a second cohort of 303 UK cases with a suspected diagnosis of MC. In 23 individuals, we found two mutations and in 86 individuals we identified a single mutation. Interestingly, 40 of the cases with a single mutation had dominant exon 8 mutations. In total 48 individuals (from 34 families) in cohort 1 and 2 were found to harbour dominant mutations (37% of mutation positive individuals, 30% of mutation positive families). In total, we have identified 23 new disease causing mutations in MC, confirming the high degree of genetic heterogeneity associated with this disease. The DNA-based strategy we have devised achieved a genetic diagnosis in 36% of individuals referred to our centre. Based on these results, we propose that exon 8 of CLCN1 is a hot-spot for dominant mutations. Our molecular expression studies of the new exon 8 mutations indicate that this region of the chloride channel has an important role in dominant negative interactions between the two chloride channel monomers. Accurate genetic counselling in MC should be based not only upon clinical features and the inheritance pattern but also on molecular genetic analysis and ideally functional expression data.

Episodic ataxia and hemiplegia caused by the 8993T->C mitochondrial DNA mutation.

The m.8993T-->C MTATP6 mutation of mitochondrial DNA (mtDNA) usually causes mitochondrial disease in childhood, but was recently described in a family with adult onset ataxia and polyneuropathy. Cytochrome c oxidase muscle histochemistry, which is the standard clinical investigation for mitochondrial disease in adults, is usually normal in patients with MTATP6 mutations. This raises the possibility that these cases have been missed in the past. We therefore studied 308 patients with unexplained ataxia and 96 patients with suspected Charcot-Marie-Tooth disease to determine whether the m.8993T-->C MTATP6 mutation is common in unexplained inherited ataxia and/or polyneuropathy. We identified a three-generation family with the m.8993T-->C mutation of mtDNA. One subject had episodic ataxia (EA) and transient hemipareses, broadening the phenotype. However, no further cases were identified in an additional cohort of 191 patients with suspected EA. In conclusion, m.8993T-->C MTATP6 should be considered in patients with unexplained ataxia, CMT or EA, but cases are uncommon.

A gene from human chromosome region 3p21 with reduced expression in small cell lung cancer.

A combination of cytogenetic and molecular studies has implicated the p21 region of human chromosome 3 as the probable site of a gene the loss of which contributes to the development of small cell lung cancer. We report here the isolation of a gene from this region which is expressed in normal lung tissue and in cell lines derived from a number of different types of tumor, but the expression of which in small cell lung cancer cell lines is undetectable by RNA blot analysis. Although the more sensitive polymerase chain reaction did detect transcripts, a novel quantitative polymerase chain reaction assay showed that their concentration in small cell lung cancer cell lines is less than 3% of that in normal lung.

Myosin rod protein: a novel thick filament component of Drosophila muscle.

Myosin rod protein (MRP), a 155 kDa protein encoded by a gene internal to the Drosophila muscle myosin heavy chain (Mhc) gene, contains the MHC rod domain, but has 77 unique N-terminal residues that exactly replace the MHC motor and light chain binding domains. Originally described as an abundant testis protein, we now demonstrate the MRP also is a major component of myofilaments in Drosophila. Specifically, the Mrp promoter directs the expression of a LacZ reporter transgene in somatic, cardiac and visceral muscles. MRP-specific antibodies detect the protein in detergent-insoluble fractions of muscle extracts and co-localize the protein with MHC to the sarcomeric A-band in immunostained muscles. Immunoblot analysis shows that in a set of adult direct flight muscles (DFM), the ratio of MRP to MHC is 1:3. Chemical cross-link and co-immunoprecipitation experiments using 0.5 M KCl-extracted thick filament proteins indicate that native MRP is a homodimer. Electron microscopy of DFM49, which has a high MRP content, shows in cross section, disordered myofilament packing and a variable thin to thick filament ratio and, in longitudinal section, severely bent thin filaments that are not well associated with thick filaments. In rigor, thick filaments from DFM49 consist of segments with cross bridges that are interspersed with smooth domains lacking cross bridges. These data indicate that MRP is a novel contractile protein that co-integrates with myosin into the thick filament, thereby changing structure and function of the sarcomere.

A genetic analysis of the alpha-glycerophosphate oxidase locus in Drosophila melanogaster.

The gene for alpha-glycerophosphate oxidase, the nuclear encoded mitochondrial enzyme of the alpha-glycerophosphate cycle (alpha GP); has been mapped in Drosophila melanogaster. Several interstitial deficiencies in region 50c-53AB of chromosome 2R were used to localize the structural gene to 52D2-5. In addition, mutations of alpha GPO were generated; alpha GPO mutants are viable yet flightless. Interactions of alpha GPO with alpha-glycerophosphate dehydrogenase (alpha GPDH), the cytoplasmic enzyme of the alpha GP cycle, were investigated through the synthesis of a series of alpha GPDHnull-alpha GPOnull double mutants. Of the six double null mutants constructed, four alpha GPDH-alpha GPO double nulls are viable and flightless. Two double mutants, however, exhibit an allelic-dependent synthetic lethal phenotype.

Splice-junction elements and intronic sequences regulate alternative splicing of the Drosophila myosin heavy chain gene transcript.

The Drosophila muscle myosin heavy chain (Mhc) gene primary transcript contains five alternatively spliced exon groups (exon 3, 7, 9, 11 and 15), each of which contains two to five mutually exclusive members. Individual muscles typically select a specific alternative exon from each group for incorporation into the processed message. We report here on the cis-regulatory mechanisms that direct the processing of alternative exons in Mhc exon 11 in individual muscles using transgenic reporter constructs, RT-PCR and directed mutagenesis. The 6.0-kilobase exon 11 domain is sufficient to direct the correct processing of exon 11 alternatives, demonstrating that the alternative splicing cis-regulatory elements are local to Mhc exon 11. Mutational analysis of Mhc exon 11 reveals that the alternative exon nonconsensus 5'-splice donors are essential for alternative splicing regulation in general, but do not specify alternative exons for inclusion in individual muscles. Rather, we show, through exon substitutions and deletion analyses, that a 360-nucleotide intronic domain precisely directs the normal processing of one exon, Mhc exon 11e, in the indirect flight muscle. These and other data indicate that alternative exons are regulated in appropriate muscles through interactions between intronic alternative splice-specificity elements, nonconsensus exon 11 splice donors and, likely, novel exon-specific alternative splicing factors.

Transposable element insertions respecify alternative exon splicing in three Drosophila myosin heavy chain mutants.

Insertions of transposable elements into the myosin heavy chain (Mhc) locus disrupt the regulation of alternative pre-mRNA splicing for multi-alternative exons in the Mhc2, Mhc3, and Mhc4 mutants in Drosophila. Sequence and expression analyses show that each inserted element introduces a strong polyadenylation signal that defines novel terminal exons, which are then differentially recognized by the alternative splicing apparatus. Mhc2 and Mhc4 have insertion elements located within intron 7c and exon 9a, respectively, and each expresses a single truncated transcript that contains an aberrant terminal exon defined by the poly(A) signal of the inserted element and the 3' acceptor of the upstream common exon. In Mhc3, a poly(A) signal inserted into Mhc intron 7d defines terminal exons using either the upstream 3' acceptor of common exon 6 or the 7d acceptor, leading to the expression of 4.1- and 1.7-kb transcripts, respectively. Acceptor selection is regulated in Mhc3 transcripts, where the 3' acceptor of common Mhc exon 6 is preferentially selected in larvae, whereas the alternative exon 7d acceptor is favored in adults. These results reflect the adult-specific use of exon 7d and suggest that the normal exon 7 alternative splicing mechanism continues to influence the selection of exon 7d in Mhc3 transcripts. Overall, transposable element-induced disruptions in alternative processing demonstrate a role for the nonconsensus 3' acceptors in Mhc exons 7 and 9 alternative splicing regulation.

Positive and negative intronic regulatory elements control muscle-specific alternative exon splicing of Drosophila myosin heavy chain transcripts.

Alternative splicing of Drosophila muscle myosin heavy chain (MHC) transcripts is precisely regulated to ensure the expression of specific MHC isoforms required for the distinctive contractile activities of physiologically specialized muscles. We have used transgenic expression analysis in combination with mutagenesis to identify cis-regulatory sequences that are required for muscle-specific splicing of exon 11, which is encoded by five alternative exons that produce alternative "converter" domains in the MHC head. Here, we report the identification of three conserved intronic elements (CIE1, -2, and -3) that control splicing of exon 11e in the indirect flight muscle (IFM). Each of these CIE elements has a distinct function: CIE1 acts as a splice repressor, while CIE2 and CIE3 behave as splice enhancers. These CIE elements function in combination with a nonconsensus splice donor to direct IFM-specific splicing of exon 11e. An additional cis-regulatory element that is essential in coordinating the muscle-specific splicing of other alternative exon 11s is identified. Therefore, multiple interacting intronic and splice donor elements establish the muscle-specific splicing of alternative exon 11s.

A model of the Drosophila melanogaster glycerol-3-phosphate oxidase-encoding gene.

We have sequenced the Drosophila melanogaster gene encoding the mitochondrial (mt) enzyme glycerol-3-phosphate oxidase (GPO; EC 1.1.99.5) that is involved in flight and alcohol metabolism. Available data suggests a simple model for this gene that includes four exons. Exon I contains a mt import signal, exon II, a transmembrane segment and an FAD-binding site, and exon IV, an iron-sulfur centre.

The human homologue of the weaver mouse gene in familial and sporadic Parkinson's disease.

The pathological hallmark of Parkinson's disease is cell death of dopaminergic neurons in the substantia nigra, resulting in striatal dopaminergic deficit and a clinical syndrome dominated by disorders of movement. The cause for this cell loss is unknown, but the possibility of a contributing genetic factor is increasingly recognized. Homozygous weaver mice, a mutant mouse strain, display progressive postnatal depletion of dopaminergic cells in the mesencephalon and have thus been proposed as an animal model for Parkinson's disease. Recently, mGIRK2, a putative G-protein inward rectifier K+ channel, has been identified as the causative gene in the weaver mouse and a homozygous mutation has been described in the H5 pore region of this channel. The human homologue of mGIRK2, KCNJ7 or hiGIRK2, has previously been isolated on chromosome 21q22.1. A possible involvement of this gene in the pathogenesis of Parkinson's disease has been discussed. To evaluate the possibility of a shared genetic defect in weaver mouse and Parkinson's disease, we analysed the H5 pore region of hiGIRK2 in familial and sporadic cases of Parkinson's disease. The sequence was normal in all cases examined, suggesting a differing aetiology of nigral cell loss in Parkinson's disease and weaver mice.

Spontaneous and induced chromosome damage in lymphocytes of medullary thyroid carcinoma patients and their family members.

Detailed chromosome analysis failed to reveal any evidence for spontaneous chromosome instability in lymphocytes from patients with multiple endocrine neoplasia, type 2 (MEN-2), whereas, with one exception, lymphocytes from MEN-2 patients were significantly more sensitive to chromosome damage by bleomycin and, to a lesser extent, MNNG than those from controls. The exceptional case suggests possible genetic heterogeneity in MEN-2.