Length Of Deletion Research Articles

Clinical outcomes of advanced NSCLC patients with different EGFR exon 19 deletion subtypes treated with first-line tyrosine kinase inhibitors: A single-center ambispective cohort study.

Clinical significance of various subtypes of epidermal growth factor receptor (EGFR) exon 19 deletion (ex19del) mutation in non-small cell lung cancer (NSCLC) remains unclear. We analyzed EGFR ex19del subtypes in NSCLC patients receiving first-line tyrosine kinase inhibitor (TKI) therapy at our center (January 2018-June 2022) and correlated them with median progression-free survival (mPFS) and median overall survival (mOS). We identified 17 different EGFR ex19del variants in 101 patients. Between the classic (E746_A750del, 64.4%) and nonclassic groups (the rest variants), no significant difference was observed in mPFS (13.5 vs. 19.3 months, p = 0.18) or mOS (44.1 vs. 77.0 months, p = 0.06). mPFS showed no significant difference between ex19del subgroups classified based on the presence of insertion (ex19delins), starting position or length of deletion. However, patients with ex19delins starting at E746 showed longer mPFS than the others (29.7 vs. 12.5 months, p = 0.04), and patients with ex19del of 15 nucleotides had shorter mOS than the others (44.1 vs. 77.0 months, p = 0.03). In multivariate analysis, ex19delins independently predicted a better PFS (HR = 0.311, p = 0.03); however, 15 nucleotide deletion was no longer associated with OS (HR = 0.181, p = 0.11). Secondary T790M mutation incidence was significantly higher in the ex19del subgroup starting at E746 than the others (64.7% vs. 30.8%, p = 0.04). Our study revealed potential differences in TKI efficacy, resistance mechanism, and prognosis of various EGFR ex19del subtypes in NSCLC, underscoring the need for precise selection of first-line therapy.

Clinical outcomes of patients with advanced NSCLC with different EGFR exon 19 deletion subtypes treated with first-line tyrosine kinase inhibitors.

e21034 Background: Epidermal growth factor receptor (EGFR) exon 19 deletion (Ex19del) in non-small cell lung cancer (NSCLC) consists of many subtypes, and their sensitivity to tyrosine kinase inhibitors (TKIs) has not been clearly defined. This study aimed to investigate the clinical outcomes of advanced NSCLC patients with different EGFR ex19del subtypes treated with different TKIs. Methods: We retrospectively analyzed stage IIIB-IV NSCLC patients with EGFR Ex19del and first-line TKIs therapy. E746_A750del was defined as classic group, and the others as non-classic group, which was further divided into subgroups according to deletion with or without insertion, starting codon or length of deletion. The primary endpoint was progression free survival (PFS). Results: There were 17 Ex19del subtypes identified in 101 patients. The frequency of E746_A750del was 64.4% (65/101), and the top three frequent non-classic subtypes were L747_T751del (6.9%, 7/101), L747_P753delinsS (5.9%, 6/101) and E746_S752delinsV (5.0%, 5/101). In all patients, the median PFS (mPFS) of first-line TKIs in classic and non-classic groups were 14.3 months (m) and 19.3 m ( p > 0.05). In non-classic group, the mPFS in deletion with insertion (delins) and without insertion subgroups were 25.3 m and 11.2 m; in deletion starting with E746 and with other codon subgroups were 29.7 m and 19.3 m; and in deletion length of 15 nucleotides (n) and of other length subgroups were 11.2 m and 25.3 m. The mPFS of these non-classic subgroups were not statistically different with that of classic group ( p > 0.05). In all patients, the mPFS of first-line therapy with the 1st, 2nd and 3rd generation TKIs were 17.4 m, 15.6 m and 20.5 m, respectively ( p＞0.05); in classic group, the mPFS were 14.3 m, 16.9 m and 20.5 m, respectively ( p＞0.05). However, in non-classic delins subgroup (29.7 m, 15.6 m and 11.9 m) and in deletion length of ≥18 n subgroup (25.3 m，12.3 m and 11.9 m), the 1st generation TKIs had an insignificantly longest mPFS ( p＞0.05). When resistant to 1st or 2nd generation TKIs, patients in classic group showed a trend of higher incidence of T790M mutation than non-classic group (72.0% vs 43.8%, p = 0.070). In multivariant analysis, pleural metastasis (PM) had interaction with Ex19del grouping, warranting stratification analysis. In patients without PM, the mPFS of first-line TKIs in classic group was significantly shorter than non-classic group (13.8 m and 29.7 m, p = 0.026). However, in patients with PM, the mPFS of classic group was numerically longer than non-classic group (16.9 m and 11.7 m, p > 0.05), with the shortest mPFS in non-classic without insertion subgroup, and in non-classic with 15 n deletion subgroup. Conclusions: In advanced NSCLC patients with EGFR Ex19del, the efficacy of first-line TKIs therapy was affected by the presence of pleural metastasis, Ex19del subtypes and different generation of TKIs.

A Novel 90-kbp Deletion of RUNX2 Associated with Cleidocranial Dysplasia.

Cleidocranial dysplasia (CCD) is a rare autosomal dominant skeletal dysplasia caused by runt-related transcription factor 2 (RUNX2) mutations. In addition to the regular missense, small or large fragment deletions are the common mutation types of RUNX2. This study aimed to find the rules of deletions in RUNX2. The clinical information of one Chinese CCD family was collected. Genomic DNA was extracted for whole-exome sequencing (WES). Bioinformatics analyzed the pathogenicity of the variants. Polymerase chain reaction (PCR) and Sanger sequencing were carried out using specific primers. RT-PCR and Q-PCR were also used to detect the mRNA level of RUNX2. The CCD studies related with deletions in RUNX2 from 1999 to 2021 from HGMD and PubMed were collected and analyzed for the relationship between the phenotypes and the length of deleted fragments. The proband presented typical CCD features, including delayed closure of cranial sutures, clavicle dysplasia, abnormal teeth. WES, PCR with specific primers and Sanger sequencing revealed a novel heterozygous 90-kbp deletion in RUNX2 (NG_008020.2 g.103671~193943), which caused a substitution (p.Asn183Ile) and premature termination (p.Asp184*). In addition, the mRNA expression of RUNX2 was decreased by 75.5% in the proband. Herein, 31 types of deletions varying from 2 bp to 800 kbp or covering the whole gene of RUNX2 were compared and the significant phenotypic difference was not found among these deletions. The CCD phenotypes were related with the final effects of RUNX2 mutation instead of the length of deletion. WES has the defects in identifying large indels, and direct PCR with specific primers and Sanger sequencing could make up for the shortcoming.

Comprehensive Analysis of Somatic Reversion Mutations in Homologous Recombination Repair (HRR) Genes in A Large Cohort of Chinese Pan-cancer Patients.

Purpose: Mutations leading to homologous recombination deficiency (HRD) increase the tumor sensitivity to platinum-based chemotherapy and PARP inhibitors. However, reversion mutations often develop conferring acquired drug resistance. There is still a lack of comprehensive investigation on HRR reversion mutations in large pan-cancer cohorts, especially in the Eastern Asian population. This study aims to characterize reversion mutations in homologous recombination repair (HRR)-related genes in a large cohort of Chinese pan-cancer patients.Methods: Sequencing data from 23,375 patients across over 17 cancer types were retrospectively analyzed for pathogenic/likely pathogenic (P/LP) germline mutations in 15 HRR genes. Somatic mutations detected in tumor or circulating cell-free DNA predicted to restore the open reading frame of the deleterious allele were subsequently identified as reversion mutations.Results: 654 cases out of 23,375 (2.8%) unselected pan-cancer patients were identified with HRR germline mutations. Secondary somatic mutations were further analyzed in their matched tumor/plasma samples. The overall frequency of reversion mutation was 1.7% (11/654). The reversion mutations occurred only in 3 out of the 15 HRR genes: BRCA1 (3.8%), BRCA2 (3.5%) and PALB2 (2.0%) from 11 patients (6 breast cancers, 1 ovarian cancer, 1 pancreatic cancer, 1 lung cancer and 2 breast and ovarian dual cancers). We identified total 25 reversion events (BRCA1, n=9; BRCA2, n=8; PALB2, n=8), including 12 pure deletions, 10 missense single nucleotide variants, 2 insertions and 1 splice site mutation. Besides, we detected microhomology length >1bp in seven out of the reversion deletions (58.3%), suggestive of microhomology-mediated end-joining (MMEJ) repair signature. Intriguingly, a positive correlation (r=0.85, p=0.001) between the length of deletion and the microhomology length was also observed. We obtained disease courses from 6/11 patients with reversion events. Four acquired reversions after the failure of the PARP inhibitor treatment. Two patients had somatic reversion mutations identified after progressing on platinum-based treatment.Conclusion: This study comprehensively depicts the prevalence and characteristics of HRR reversion mutation of germline mutations in an unselected Chinese pan-cancer cohort. The reversion mutations predominantly occurred in BRCA1, BRCA2 and PALB2. The results revealed that reversion mutations frequently occurred after resistance to platinum-based chemotherapy and/or PARP inhibitor. Our study provides insight into the underlying mechanism of drug resistance in HRD tumors and suggests that monitoring HRR mutation status along the disease course could be beneficial especially for informing resistance mechanisms and guiding subsequent therapies.

Single-Strand Breaks at Genome Initiate FLT3-ITD Formation

Single-Strand Breaks at Genome Initiate FLT3-ITD Formation

Genetic Modifiers of the Spinal Muscular Atrophy Phenotype

Proximal spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disease caused by homozygous deletion in the seventh exon of the SMN1 gene. The aim of this work is to analyze the association of the allelic polymorphism of telomeric genes SMN1 and NAIP and the centromeric gene SMN2 of the 5q13 region with the clinical phenotype of SMA. It was shown that the homozygous genotype, which contains a telomeric deletion, covering both SMN1 and NAIP, is significantly more often observed in patients with the most severe type of SMA. Three or more copies of SMN2 are associated with a milder phenotype; the number of SMN2 copies affects the SMA phenotype more heavily than the length of the telomeric deletion. It was shown that one SMN2 copy is significantly more frequent than three or more copies of this gene in SMA-patients with homozygous deletion of SMN1 and NAIP. This fact may indicate the presence of a large deletion of all the three studied genes in SMA genotypes associated with the most severe type of SMA. It is noted that congenital SMA (type 0) is significantly less common in female patients, which may indicate the presence of SMA modifier genes on the X-chromosome.

The preS deletion of hepatitis B virus (HBV) is associated with liver fibrosis progression in patients with chronic HBV infection.

Limited data are available regarding the association of hepatitis B virus (HBV) mutations with liver fibrosis in HBV infection. The study aimed to clarify whether HBV preS deletion mutation is associated with liver fibrosis progression. A total of 469 patients were enrolled, including 324 with chronic hepatitis B (CHB), 28 with HBV-related compensated liver cirrhosis (LC), and 117 with HBV-related decompensated LC. All CHB and compensated LC patients received liver biopsy. Fibrosis grade was assessed using METAVIR score. HBV preS deletion was determined by direct sequencing and verified by clonal sequencing. Overall preS deletion was detected in 12.6% (59/469) patients, specifically, in 7.51% (13/173), 10.60% (16/151), and 20.69% (30/145) of patients with no-to-mild liver fibrosis (F0-1), moderate-to-severe liver fibrosis (F2-3), and cirrhosis (F4), respectively (p < 0.01). Patients with preS-deleted HBV had lower serum HBV DNA and albumin levels compared to patients with wild-type HBV. The median length of preS deletion was 39-base pairs (bp) (3-204bp) and the deletion most frequently emerged in preS2 initial region. Multivariate analysis identified the preS2 deletion rather than preS1 deletion to be an independent risk factor of significant fibrosis, i.e., METAVIR F ≥ 2 (p = 0.007). In addition, preS-deleted viral sequences were detected in the pool of intrahepatic HBV covalently closed circular DNA. HBV preS deletion is positively associated with liver fibrosis progression in chronic HBV-infected patients. HBV preS2 deletion may serve as a warning indicator for liver fibrosis progression.

Mutational analysis of foot and mouth disease virus nonstructural polyprotein 3AB-coding region to design a negative marker virus

Inactivated purified whole virus vaccines are used for control of foot and mouth disease (FMD). ELISAs detecting antibodies to the nonstructural proteins (NSP), a marker of infection, are primarily used to differentiate FMD virus (FMDV) infected from vaccinated animals (DIVA). However, such DIVA assays have a limitation to their specificity since residual NSPs present in the relatively impure vaccines are suspected to induce an NSP-antibody response in the repeatedly vaccinated animals. Epitope-deleted negative marker vaccine strategy seems to have an advantage over the conventional vaccines in identifying the infected animals with accuracy. NSP 3AB contains an abundance of immunodominant B-cell epitopes of diagnostic importance. This study addresses the feasibility of producing 3AB-truncated FMDV mutant as a potential negative marker vaccine candidate. An infectious cDNA clone of FMDV serotype Asia 1 strain was used to engineer an array of deletion mutations in the established antigenic domain of 3AB. The maximum length of deletion tolerated by the virus was found to be restricted to amino acid residues 87–144 in the C-terminal half of 3A protein along with deletion of the first two copies of 3B peptide. The 3AB-truncated marker virus (Asia 1 IND 491/1997Δ3A87-1443B1,2+FLAG) demonstrated infectivity titres comparable to that of the parental virus in BHK-21 (log10 7.42 TCID50/ml) and LFBK-αVβ6 (log10 8.30 TCID50/ml) cell monolayer culture. The protein fragment corresponding to the viable deletion in the 3AB region was expressed in a prokaryotic system to standardize a companion assay (3A87-1533B1,2 I-ELISA) for the negative marker virus which showed reasonably high diagnostic sensitivity (96.9%) and specificity (100% for naïve and 97.1% for uninfected vaccinated samples). The marker virus and its companion ELISA designed in this study provide a basis to devise a marker vaccine strategy for FMD control.

Y chromosomal deletion pattern in Koreans inhabiting Jeju Island

Mutations occur in Y chromosome genes similar to autosomal genes. However, unlike autosomal genes, Y chromosome genes do not undergo recombination, which produce distinctive characteristics and distribution patterns in different geographic regions. Therefore, detailed analysis of mutations of Y chromosome genes might provide information for personal identification or analysis of phylogenetic history. In Y-STR (short tandem repeat) analysis tests on 668 habitants of Jeju Island, the largest island in the Korean peninsula located apart from the mainland, a deletion at DYS448 was found in 10 samples. The length of deletion was estimated by confirming specific Sequence Tagged Site (STS) markers ranging from G66018 to sY1201. Patterns found were similar to those of the Kalmyks, a tribe that has had strong social and genetic influences in Jeju Island in the past. Historically from 1273 on, Jeju Island was governed by Mongolian for about one hundred years. The results of this study suggest such historical aspects affected the genetic composition of people living in Jeju Island. Furthermore, previous reports showed that Y chromosomal deletions and region specific Y chromosomal mutations depended on regional differences. This study may be useful for a better understanding of the genetic structure of Jeju habitants as well as Korean population for the purpose of forensic practice and population genetics.

The activation peptide of coagulation factor XIII is vital for its expression and stability

The human activation peptide of factor XIII (AP-FXIII) comprises the first 37 amino acids of the N-terminus and holds the FXIII in an inactive state. FXIII is activated either proteolytically by cleavage of AP-FXIII by thrombin, or non-proteolytically by high calcium concentrations. To investigate the role of AP-FXIII in the expression and stability of FXIII. We cloned 13 FXIII variants with progressivetruncations of AP-FXIII from the N-terminus (delN-FXIII-A), expressed them in mammalian cells, and measured their thermostability, activation, and transglutaminase activity. We also used in silico calculations to analyze the stability of hypothetical delN-FXIII dimers and to identify crucial motifs within AP-FXIII. Variants with deletions longer than the first 10 amino acids and an R11Q point mutant were not expressed as proteins. In silico calculations indicated that the sequence (8) FGGR(12) R plays a substantial role in intersubunit interactions in FXIII-A2 homodimers. In agreement with this prediction, the temperature stability of delN-FXIII variants decreased with increasing length of deletion. These results may suggest a role of the N-terminus of AP-FXIII in dimer stability. Substantial sequence homology was found among activation peptides of vertebrate and even invertebrate (crustacean) FXIII-A orthologs, which further supports our conclusion. We conclude that deletion of 11 or more N-terminal amino acids disrupts intersubunit interactions, which may prevent FXIII-A2 homodimer formation. Therefore, AP-FXIII plays an important role in the stability of the FXIII-A2 dimer.

Novel, closely related, white spot syndrome virus (WSSV) genotypes from Madagascar, Mozambique and the Kingdom of Saudi Arabia

White spot syndrome virus (WSSV) is highly pathogenic to penaeid shrimp and has caused significant economic losses in the aquaculture industry around the world. During 2010 to 2012, WSSV caused severe mortalities in cultured penaeid shrimp in Saudi Arabia, Mozambique and Madagascar. To investigate the origins of these WSSV, we performed genotyping analyses at 5 loci: the 3 open reading frames (ORFs) 125, 94 and 75, each containing a variable number of tandem repeats (VNTR), and deletions in the 2 variable regions, VR14/15 and VR23/24. We categorized the WSSV genotype as {N125, N94, N75, ΔX14/15, ΔX23/24} where N is the number of repeat units in a specific ORF and ΔX is the length (base pair) of deletion within the variable region. We detected 4 WSSV genotypes, which were characterized by a full-length deletion in ORF94/95, a relatively small ORF75 and one specific deletion length in each variable region. There are 2 closely related genotypes in these 3 countries: {6125, del94, 375, Δ595014/15, Δ1097123/24} and {7125, del94, 375, Δ595014/15, Δ1097123/24}, where del is the full-length ORF deletion. In Saudi Arabia, 2 other related types of WSSV were also found: {6125, 794, 375, Δ595014/15, Δ1097123/24} and {8125, 1394, 375, Δ595014/15, Δ1097123/24}. The identical patterns of 3 loci in these 4 types indicate that they have a common lineage, and this suggests that the WSSV epidemics in these 3 countries were from a common source, possibly the environment.

Correlation of Vernalization Loci VRN-H1 and VRN-H2 and Growth Habit in Barley Germplasm

Vernalization requirement is a key component in determining the overall fitness of developmental patterns of barley to its environment. We have used previously reported markers and spring-sown growth habit nursery to characterize the genotypes of barley germplasm in an applied barley breeding ground to establish a baseline of information required to understand the relationship between adaptation of autumn-sown barley germplasm in diverse regions with warm (W), moderate (M), or cold climates (C). This study revealed that twenty entries were detected with the presence of the vernalization critical region in VRN-H1 locus and complete presence of the three geneclusters ZCCT-Ha, -Hb, and -Hc in VRN-H2 locus represented as genotype vrn-H1/Vrn-H2 (V1w/V2w). Of these genotypes, 17 entries showed winter growth habit whereas the remaining three revealed facultative growth habit indicating reduced vernalization requirements possibly due to VRN-H3 and photoperiod sensitivity loci as compared to the landmark winter growth habit entries in this group. Twenty-four entries were detected with the lack of vernalization critical region in VRN-H1 locus but complete presence of the three geneclusters ZCCT-Ha, -Hb, and -Hc in VRN-H2 locus represented as genotype Vrn-H1/Vrn-H2 (V1s/V2w). However, only half of these germplasms were identified with spring growth habit in spring-sown nursery, and the rest of the germplasms in this group revealed facultative growth habits due to possible variation in the length of deletion in VRN-H1. Four germplasms showed vernalization insensitive phenotype due to the lack of a functional ZCCT-Ha and/or ZCCT-Hb alleles in VRN-H2 and the deletion in the vernalization critical region of VRN-H1. These germplasms revealed acomplete spring type growth habit. Only one entry showed reduced vernalization requirement solely due to the deletion in functional ZCCT-Hb allele in VRN-H2 and not due to the deletion in the vernalization critical region of VRN-H1.

Evaluation of trace mineral source and preharvest deletion of trace minerals from finishing diets for pigs on growth performance, carcass characteristics, and pork quality1,2,3

Weanling crossbred pigs (Sus scrofa; 72 barrows and 72 gilts; BW = 7.4 ± 1.1 kg) were used to evaluate dietary supplemental trace mineral (Cu, Fe, Mn, and Zn) source (inorganic vs. organic) and deletion (0, 2, 4, and 6 wk preharvest) on growth performance, carcass characteristics, and pork quality. Pigs were blocked by BW, ancestry, and sex, and randomly allotted to 24 pens, and fed a diet containing either inorganic or organic trace minerals supplemented at the 1998 NRC requirement estimates for each of 5 BW phases from 7 to 120 kg (equivalent to 14, 14, 42, 28, and 42-d periods, respectively). Two pigs were removed from each pen at the end of Phase IV (BW = 82.6 ± 6.0 kg), and 2 other pigs were removed at the end of Phase V (BW = 128.0 ± 8.3 kg) for collection of various tissues and for determination of carcass characteristics and pork quality. On d 1, 15, and 29 of Phase V, 3 pens within each source of minerals were switched to a common diet without supplemental trace minerals, whereas the remaining 3 pens within each source of minerals were fed diets containing trace minerals throughout the Phase V period. This resulted in 4 groups within each mineral treatment, in which trace mineral supplementation was deleted for 6, 4, 2, or 0 wk of Phase V. Trace mineral source (inorganic vs. organic) did not affect ADG, ADFI, and G:F (773 vs. 778 g/d, 1,680 vs. 1,708 g/d, and 461 vs. 456 g/kg, respectively) during the first 4 phases. During the mineral deletion period, ADG and G:F were not affected by the duration of trace mineral deletion, but ADFI increased when trace minerals were removed from the diet for 6 wk (6 vs. 0 wk, 3,393 vs. 3,163 g/d; P = 0.05). Hot carcass weight, cold carcass weight, carcass shrink, dressing percentage, LM area, 10th rib and midline average backfat, and carcass fat-free lean weight and percentage were not affected (P > 0.10) by the source of mineral or length of mineral deletion, but carcass length tended to decrease (P = 0.09) when time of trace mineral deletion increased. Increasing mineral deletion from 0 to 6 wk tended to reduce linearly (P = 0.08) Hunter a* scores on the day of carcass processing (24 h after slaughter), as well as 2 d after processing, and Hunter b* scores on d 2 and d 6 after processing. Results of this experiment indicate that use of organic trace minerals, rather than inorganic trace minerals, did not influence pig growth performance or carcass characteristics and quality; however, deletion of minerals during the last 6 wk before harvest increased ADFI and affected drip loss, some color scores of the LM, and carcass length.

A child with a deletion in the monocarboxylate transporter 8 gene: 7-year follow-up and effects of thyroid hormone treatment

The monocarboxylate transporter 8 (MCT8; SLC16A2) has a pivotal role in neuronal triiodothyronine (T(3)) uptake. Mutations of this transporter determine a distinct X-linked psychomotor retardation syndrome (Allan-Herndon-Dudley syndrome (AHDS)) that is attributed to disturbed thyroid hormone levels, especially elevated T(3) levels. We describe the genetic analysis of the MCT8 gene in a patient suspected for AHDS and the clinical and endocrine effects of L-thyroxine (LT(4)) or liothyronine (LT(3)) treatment intending to overcome the T(3) uptake resistance through alternative transporters. The six exons of the MCT8 gene were amplified individually by PCR. As multiple exons were missing, the length of the X-chromosomal deletion was determined by a dense SNP array, followed by PCR-based fine mapping to define the exact borders of the deleted segment. The clinical and endocrine data of the patient during 6.5 years of LT(4) treatment and two periods (3 months each) of low- and high-dose LT(3) were evaluated. A partial deletion of the MCT8 gene (comprising five of six exons) was detected, confirming the suspected AHDS. MCT8 dysfunction was associated with partial resistance to T(3) at the hypothalamus and pituitary level, with normal responsiveness at the peripheral organs (liver and cardiovascular system). Thyroid hormone administration had no beneficial effect on the neurological status of the patient. We identified a 70 kb deletion encompassing exons 2-6 of the MCT8 gene in our AHDS patient. Both LT(4) and LT(3) administration had no therapeutic effect. Alternatively, treatment of AHDS patients with thyroid hormone analogs should be considered.

Association of mutation types and distribution characteristics of dystrophin gene with clinical symptoms in Chinese population

Duchenne muscular dystrophy (DMD) is X-linked disorder caused by mutations in the dystrophin gene. To investigate mutation types and distribution characteristics of dystrophin gene in Chinese DMD patients, we used Multiplex Ligation-Dependent Probe Amplification (MLPA) to analyze the dystrophin gene in 720 DMD patients, their mothers, and 20 normal adult males. Results showed that detection rate was 64.9% (467/720) in all the patients, gene deletion rate was 54.3% (391/720), and gene duplication rate was 10.6% (76/720). The rate of deletion mutant occurred in Exon 45-54 was 71.9% (281/391) in all gene deletion patients; meanwhile, the rate of gene duplication occurred in Exon 1-40 was 82.9% (63/76) in all gene duplication ones. In all the patients with gene deletion and duplication, the rate of DMD and IMD was 90.6% (423/467), and BMD, 9.4% (44/467). This indicates that the main reason of duchenne muscular dystrophy is dystrophin gene deletion mutation, which would occur in any gene unevenly with hot spots of mutation. The location and fragment length of gene deletion and duplication cannot decide the severity of clinical symptoms directly.

Diagnosis and fine localization of deletion region in Wolf- Hirschhorn syndrome patients

Background Wolf-Hirschhorn syndrome (WHS) results from the partial deletion of 4p. This study aimed to identify and finemap the chromosome deletion regions of Chinese children with Wolf-Hirschhorn syndrome among the developmental delay/mental retardation (DD/MR) patients. Methods We analyzed the relationship of phenotype and genotype. Inclusion criteria were: moderate to severe DD/MR, no definite perinatal brain injury, and no trauma, toxication, hypoxia, infection of central nervous system; routine karyotyping was normal, no evidence of typical inherited metabolic disorder or specific neurodegenerative disorders from cranial neuro-imaging and blood/urinary metabolic diseases screening; no mutation of FMR1 in male patients, no typical clinical manifestation of Rett syndrome in female patients. Multiplex ligation-dependent probe amplification (MLPA) and Affymetrix genome-wide human SNP array 6.0 assays were applied to accurately define the exact size of subtelomeric aberration region of four WHS patients. Results All four WHS patients presented with severe DD, hypotonia and microcephaly, failure to thrive, 3/4 patients with typical facial features and seizures, 2/4 patients with congenital heart defects and cleft lip/palate, 1/4 patients with other malformations. The length of the deletions ranged from 3.3 Mb to 9.8 Mb. Two of four patients had “classic” WHS, 1/4 patients had “mild”-to-“classic” WHS, and 1/4 patients had “mild” WHS. Conclusions WHS patients in China appear to be consistent with those previously reported. The prevalence of signs and symptoms, distribution of cases between “mild” and “classic” WHS, and the correlation between length of deletion and severity of disease of these patients were all similar to those of the patients from other populations.

A Novel Mechanism of P-type ATPase Autoinhibition Involving Both Termini of the Protein

The activity of many P-type ATPases is found to be regulated by interacting proteins or autoinhibitory elements located in N- or C-terminal extensions. An extended C terminus of fungal and plant P-type plasma membrane H(+)-ATPases has long been recognized to be part of a regulatory apparatus involving an autoinhibitory domain. Here we demonstrate that both the N and the C termini of the plant plasma membrane H(+)-ATPase are directly involved in controlling the pump activity state and that N-terminal displacements are coupled to secondary modifications taking place at the C-terminal end. This identifies the first group of P-type ATPases for which both ends of the polypeptide chain constitute regulatory domains, which together contribute to the autoinhibitory apparatus. This suggests an intricate mechanism of cis-regulation with both termini of the protein communicating to obtain the necessary control of the enzyme activity state.

Is the 1p/19q deletion a diagnostic marker of oligodendrogliomas?

The diagnosis and classification of diffusely infiltrative gliomas are based on their histopathological appearance; however, histopathological delineation of diffuse gliomas can be difficult because of vague and subjective histopathological criteria. Combined loss of chromosome arms 1p and 19q (denoted as 1p−/19q−) has proven to be a powerful predictor of chemotherapeutic response and survival in oligodendrogliomas. We undertook retrospective and prospective studies of brain tumor patients originally diagnosed as oligodendrogliomas or oligoastrocytomas patients followed at our institution using molecular genetic techniques. Fluorescence in situ hybridization using probes specific for chromosomes 1 and 19 was performed on 22 paraffin-embedded tissues retrospectively; 15 touch-preparation smear samples were studied prospectively; and loss of heterozygosity (LOH) screening was performed on 11 samples with microsatellite markers specific to chromosome 1 and chromosome 19. Of the 37 cases, 24 had 1p−/19q−, 1 case had 1p− only, 2 cases had 19q− only, and 10 cases had no deletion. The length of the largest deletion was mapped between markers D1S2795 (1p36.31 locus) and D1S2722 (1p34.2 locus) and between markers D19S416 (19q13.11 locus) and D19S397 (19q13.14 locus), using LOH. All of the pure oligodendrogliomas ( n = 7) harbored 1p−/19q−. In light of previous findings, the 1p−/19q− combination appears to be an objective diagnosis marker of classic oligodendrogliomas, one that can be used, in combination with histological examination, to improve the diagnosis of oligodendroglioma. Fluorescence in situ hybridization on touch preparations is a simple way to obtain information on 1p−/19q− in 24 hours.

Genotype and phenotype analyses in 136 patients with single large-scale mitochondrial DNA deletions

We examined 136 patients with mitochondrial DNA (mtDNA) deletion. Clinical diagnoses included chronic progressive external ophthalmoplegia (94 patients); Kearns-Sayre syndrome (KSS; 33 patients); Pearson's marrow-pancreas syndrome (six patients); and Leigh syndrome, Reye-like syndrome, and mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes (one patient). The length and location of deletion were highly variable. Only one patient had deletion within the so-called shorter arc between the two origins of mtDNA replication. The length of deletion and the number of deleted transfer ribonucleic acid (tRNAs) showed a significant relationship with age at onset. Furthermore, KSS patients had longer and larger numbers of deleted tRNAs, which could be risk factors for the systemic involvement of single mtDNA deletion diseases. We found 81 patterns of deletion. Direct repeats of 4 bp or longer flanking the breakpoints were found in 96 patients (70.5%) and those of 10 bp or longer in 49 patients (36.0%). We found two other common deletions besides the most common deletion (34 patients: 25.0%): the 2,310-bp deletion from nt 12113 to nt 14421 (11 patients: 8.0%) and the 7,664-bp deletion from nt 6330 to nt 13993 (ten patients: 7.3%). These deletions had incomplete direct repeats longer than 13 bp with one base mismatch.

Hydrophilic Residues 526KNDAAD531 in the Flexible C-terminal Region of the Chaperonin GroEL Are Critical for Substrate Protein Folding within the Central Cavity

The final 23 residues in the C-terminal region of Escherichia coli GroEL are invisible in crystallographic analyses due to high flexibility. To probe the functional role of these residues in the chaperonin mechanism, we generated and characterized C-terminal truncated, double ring, and single ring mutants of GroEL. The ability to assist the refolding of substrate proteins rhodanese and malate dehydrogenase decreased suddenly when 23 amino acids were truncated, indicating that a sudden change in the environment within the central cavity had occurred. From further experiments and analyses of the hydropathy of the C-terminal region, we focused on the hydrophilicity of the sequence region (26 KNDAAD 531 and generated two GroEL mutants where these residues were changed to a neutral hydropathy sequence (526 GGGAAG 531) and a hydrophobic sequence (526 IGIAAI 531), respectively. Very interestingly, the two mutants were found to be defective in function both in vitro and in vivo. Deterioration of function was not observed in mutants where this region was replaced by a scrambled (526 NKADDA 531) or homologous (526 RQEGGE 531) sequence, indicating that the hydrophilicity of this sequence was important. These results highlight the importance of the hydrophilic nature of 526 KNDAAD 531 residues in the flexible C-terminal region for proper protein folding within the central cavity of GroEL.