Carlos Córdova-Fletes

Oxidative stress modulation in hepatitis C virus infected cells

Hepatitis C virus (HCV) replication is associated with the endoplasmic reticulum, where the virus can induce cellular stress. Oxidative cell damage plays an important role in HCV physiopathology. Oxidative stress is triggered when the concentration of oxygen species in the extracellular or intracellular environment exceeds antioxidant defenses. Cells are protected and modulate oxidative stress through the interplay of intracellular antioxidant agents, mainly glutathione system (GSH) and thioredoxin; and antioxidant enzyme systems such as superoxide dismutase, catalase, GSH peroxidase, and heme oxygenase-1. Also, the use of natural and synthetic antioxidants (vitamin C and E, N-acetylcysteine, glycyrrhizin, polyenylphosphatidyl choline, mitoquinone, quercetin, S-adenosylmethionine and silymarin) has already shown promising results as co-adjuvants in HCV therapy. Despite all the available information, it is not known how different agents with antiviral activity can interfere with the modulation of the cell redox state induced by HCV and decrease viral replication. This review describes an evidence-based consensus on molecular mechanisms involved in HCV replication and their relationship with cell damage induced by oxidative stress generated by the virus itself and cell antiviral machinery. It also describes some molecules that modify the levels of oxidative stress in HCV-infected cells.

CDKL5 truncation due to a t(X;2)(p22.1;p25.3) in a girl with X-linked infantile spasm syndrome

To the Editor : X-linked infantile spasm syndrome (ISSX; OMIM 308350) occurs in 2–5 of 10,000 newborns and is characterized by early onset seizures, hypsarrhythmia, and severe mental retardation. Three girls with ISSX and an X;autosome translocation that disrupted the CDKL5 gene have been reported (1–3), whereas ∼50 different CDKL5 mutations have been found in >60 clinically similar patients with normal chromosomes (4–15). We report a girl with severe ISSX and a t(X;2) disrupting the CDKL5 gene in intron 3 and fusing CDKL5 and RPS7 genes. The proposita (Fig. 1a,b) was born to a 35year-old mother and a 45-year-old father. Since the age of 1 month, she presented myoclonic seizures with cyanosis that were partially controlled with several drugs. Her psychomotor development has been profoundly delayed. At the age of 2 years, she exhibited generalized hypotonia, lack of eye contact, poor pupilar reflex, weight of 7300 g [−5.5 standard deviation (SD)], length of 79 cm (−4.5 SD), occipital-frontal circumference (OFC) of 45 cm (−3 SD/−2.5 SD), narrow forehead, frontal hypertrichosis, synophrys, upslanted palpebral fissures, broad nasal bridge, small nose and diminished muscle bulk. Evoked visual and auditory potentials were consistent with central blindness and visual pathway immaturity, but showed normal hearing. Cranial computed tomography scan revealed fronto-temporal cortical atrophy and corpus callosum hypoplasia (Fig. 1c). Repeated electroencephalograms disclosed generalized paroxysmal activity and disorganization. The patient and her mother were karyotyped on GTG-banded chromosomes; the former had a 46,X,t(X;2)(p22.1;p25.3) karyotype assumed to be de novo (Fig. 1d) as the mother was 46,XX and the unlikelihood of the father (not sampled) being a carrier. A preferential inactivation of the normal X was found in 30 RBG-banded metaphases and confirmed by the human androgen receptor assay (HUMARA) (16) that also revealed a maternal origin for this chromosome (data not shown); i.e. the der(X) was the paternal homologue. Genescan analysis confirmed a ∼90% inactivation of the maternal allele (MA) (Fig. 1e). The X-chromosome inactivation in the mother was at random. Fluorescence in situ hybridization (FISH) with clones RP11-558P14 (AL109798) and RP1245G19 (Z92542) revealed that the X-chromosome breakpoint disrupted CDKL5 (Fig. 2a). Reverse transcriptase-polymerase chain reaction (RT-PCR) on patient RNA showed products with CDKL5 primer pairs spanning exons 9–12 (Fig. 2c) and exons 13–18 (data not shown). No product could be obtained using primers located in exons 2 and 8 (Fig. 2c). These results let us hypothesize that the chromosome 2 breakpoint mapped also within a gene and the rearrangement had produced fusion genes. To test this, we have fine mapped the breakpoints by array painting as described previously (17, 18). The results indicated that on der(X) the breakpoint lies within CDKL5 intron 3 (chrX:18,447,901-18,460,135) (UCSC, hg18) and on der(2) in a region that contained the 5′ segment, including exons 1 and 2, of the RPS7 gene (chr2:3,599,632-3,601,126). RT-PCR with CDKL5 exon 2 forward primer and RPS7 reverse primers for amplifying fusion transcripts from the der(2) resulted in products containing CDKL5 exons 2 and 3 spliced to chromosome 2 position 3,600,514 within a single exon EST (expressed sequence tags) (DA710611), followed by RPS7 exons 1–6 (shown in Fig. 2d). This result let us to conclude that the single exon EST, or at least a part of it, presents an extension of the RPS7 5′UTR. Therefore, the ORF of RPS7, which codes for a ribosomal protein of the small subunit, is most likely not affected by the translocation but the CDKL5–RPS7 fusion gene is very likely transcribed from the CDKL5 promoter. For der(X), a similar approach for amplifying

A de novo t(10;19)(q22.3;q13.33) leads to ZMIZ1/PRR12 reciprocal fusion transcripts in a girl with intellectual disability and neuropsychiatric alterations.

We report a girl with intellectual disability (ID), neuropsychiatric alterations, and a de novo balanced t(10;19)(q22.3;q13.33) translocation. After chromosome sorting, fine mapping of breakpoints by array painting disclosed disruptions of the zinc finger, MIZ-type containing 1 (ZMIZ1) (on chr10) and proline-rich 12 (PRR12) (on chr19) genes. cDNA analyses revealed that the translocation resulted in gene fusions. The resulting hybrid transcripts predict mRNA decay or, if translated, formation of truncated proteins, both due to frameshifts that introduced premature stop codons. Though other molecular mechanisms may be operating, these results suggest that haploinsufficiency of one or both genes accounts for the patient’s phenotype. ZMIZ1 is highly expressed in the brain, and its protein product appears to interact with neuron-specific chromatin remodeling complex (nBAF) and activator protein 1 (AP-1) complexes which play a role regulating the activity of genes essential for normal synapse and dendrite growth/behavior. Strikingly, the patient’s phenotype overlaps with phenotypes caused by mutations in SMARCA4 (BRG1), an nBAF subunit presumably interacting with ZMIZ1 in brain cells as suggested by our results of coimmunoprecipitation in the mouse brain. PRR12 is also expressed in the brain, and its protein product possesses domains and residues thought to be related in formation of large protein complexes and chromatin remodeling. Our observation from E15 mouse brain cells that a Prr12 isoform was confined to nucleus suggests a role as a transcription nuclear cofactor likely involved in neuronal development. Moreover, a pilot transcriptome analysis from t(10;19) lymphoblastoid cell line suggests dysregulation of genes linked to neurodevelopment processes/neuronal communication (e.g., NRCAM) most likely induced by altered PRR12. This case represents the first constitutional balanced translocation disrupting and fusing both genes and provides clues for the potential function and effects of these in the central nervous system.

Delineation of a de novo 7q21.3q31.1 Deletion by CGH-SNP Arrays in a Girl with Multiple Congenital Anomalies Including Severe Glaucoma.

In this study, we present a female patient with a constitutional de novo deletion in 7q21.3q31.1 as determined by G-banding and CGH-SNP arrays. She exhibited, among other features, psychomotor retardation, congenital severe bilateral glaucoma, a cleft palate, and heart defect. Microarray assay disclosed a deleted 12.5-Mb region roughly 88 kb downstream the ectrodactyly critical region; thus, the patients final karyotype was 46,XX.arr 7q21.3q31.1(96,742,140- 109,246,085)×1 dn. This girl represents the fourth patient described so far with congenital glaucoma and a deletion encompassing or overlapping the 7q21.3q31.1 region, and confirms the presence of a locus or loci related to such a clinical feature. According to our results, the proneness to ocular defects secondary to 7q intermediate deletions could be caused by co-deletion of TAC1, HBP1, and a small cluster of cytochrome P450 genes (subfamily 3A). This conclusion is supported by their functional roles and expression locations as well as because TAC1 is related to the functional pathway of the MYOC gene whose mutations are linked to glaucoma. Moreover, given that this girl is clinically reminiscent of several phenotypes related to diverse deletions within 7q21q32, our results and observations offer a general overview of the gene content of deletions/phenotypes overlapping 7q21.3q31.1 and confirm that loci distal to DLX genes including the CUX1 gene and potential regulatory elements downstream from DLX5 are unrelated to ectrodactyly.

Complex 9p Rearrangement in an XY Patient With Ambiguous Genitalia and Features of Both 9p Duplication and Deletion

Complex 9p Rearrangement in an XY Patient With Ambiguous Genitalia and Features of Both 9p Duplication and Deletion Vivian Alejandra Neira, Carlos C ordova-Fletes,* Yohann Grondin, Azubel Ramirez-Velazco, Luis E. Figuera, Roc io Ort iz-L opez, and Michela Barbaro Divisi on de Gen etica, Centro de Investigaci on Biom edica de Occidente, CMNO-IMSS, Guadalajara, M exico Doctorado en Gen etica Humana, Universidad de Guadalajara, Guadalajara, Jalisco, M exico Unidad de Biolog ia Molecular, Gen omica y Secuenciaci on, Centro de Investigaci on y Desarrollo en Ciencias de la Salud, Universidad Aut onoma de Nuevo Le on, Monterrey, Nuevo Le on, M exico Facultad deMedicina, DepartamentodeBioqu imicayMedicinaMolecular, UniversidadAut onomadeNuevoLe on,Monterrey, NuevoLe on,M exico Facultad de Ciencias Biol ogicas, Universidad Aut onoma de Nuevo Le on, Monterrey, Nuevo Le on, M exico Department of Molecular Medicine and Surgery, Center for Molecular Medicine, Karolinska University Hospital, Stockholm, Sweden

A de novo sSMC(22) Characterized by High-Resolution Arrays in a Girl with Cat-Eye Syndrome without Coloboma.

Cat-eye syndrome (CES) results from trisomy or tetrasomy of proximal 22q originated by a small supernumerary marker chromosome (sSMC). Two critical regions for the major clinical features of CES (CESCRs) have been suggested; however, CES clinical presentation often does not correlate with the sSMC genetic content. We report here a CES girl without coloboma and carrier of a de novo type I sSMC(22) as determined by G- and C-banding, NOR staining and microarrays. This sSMC included 6 distal genes outside the original CESCR and led to a tetrasomy for 22q11.1–22q11.21. The patient’s final karyotype was 47,XX,+psu dic(22)(q11.21).arr 22q11.1q11.21(15,250,000–17,035,860)×4 dn. The amplified region outside of CESCR included some genes that may be related to neurologic, heart and renal abnormalities. Conversely, even though the amplification included the CECR2 gene, a major candidate for eye features, there was no coloboma in the patient. The genetic delineation of the present sSMC further strengthens that the CES clinical presentation does not fit completely with the duplicated genetic content and that CES is actually a genomic disorder. Furthermore, since we observed no mosaicism, we believe that other mechanisms might be behind the variability of CES phenotypes as well, mainly those related with functional interactions among amplified genes.

De novo MECP2 disomy in a Mexican male carrying a supernumerary marker chromosome and no typical Lubs syndrome features.

Xq28 duplication, including the MECP2 gene, is among the most frequently identified Xq subtelomeric rearrangements. The resulting clinical phenotype is named Lubs syndrome and mainly consists of intellectual disability, congenital hypotonia, absent speech, recurrent infections, and seizures. Here we report a Mexican male patient carrying a supernumerary marker chromosome with de novo Xq28 gain. By MLPA, duplication of MECP2, GDI1, and SLC6A8 was found and a subsequent a-CGH analysis demonstrated that the gain spanned ~2.1Mb. Despite gain of the MECP2 gene, the features of this patient do not evoke Lubs syndrome. Probably the mosaicism of the supernumerary marker chromosome is modifying the phenotype in this patient.

Exome sequencing reveals three homozygous missense variants in SNRPA in two sisters with syndromic intellectual disability.

Splicing‐related gene mutations might affect the expression of a single gene or multiple genes and cause clinically heterogeneous diseases. With the advent of next‐generation sequencing, several splicing gene mutations have been exposed, yet most major spliceosome genes have no reports of germline mutations and therefore, their effects are largely unknown. We describe the previously unreported concurrence of intellectual disability, short stature, poor speech, and minor craniofacial and hand anomalies in 2 female siblings with 3 homozygous missense variants in SNRPA (a component of the U1 small nuclear ribonucleoprotein complex) characterized by homozygosity mapping and whole exome sequencing. Combined, c.97A>G, c.98T>C, and c.100T>A, in exon 2 of SNRPA lead to p.Ile33Ala and p.Phe34Ile exchanges, which were predicted in silico to be deleterious. Although both patients exhibited some clinical features seen in other spliceosomal disorders, their complete clinical phenotype appears to be rather uncommon, a finding that may further support the notion that mutations in components of the major spliceosome do not strictly lead to the same syndromes/phenotypes.

De Novo San Luis Valley Syndrome-like der(8) Chromosome With a Concomitant dup(8p22) in a Mexican Girl

The rec(8)dup(8q)inv(8)(p23.1q22.1) chromosome associated with San Luis Valley Syndrome (SLVS OMIM 179613) is usually diagnosed in Hispanic patients from the USA Southwest where a founder carrier Spaniard lived around 1800 [1, 2]. This rec(8) has an 8q duplication of 47.90 Mb and an 8p deletion of 11.65 Mb [3, 4]. Excluding two de novo rec(8)dup q chromosomes characterized only by G-bands and included in a recent compilation [5], cytogenomic analyses identified nine comparable de novo der(8)dup q/del p chromosomes with or without a simultaneous 8p gain. We describe a Mexican mestizo girl with a de novo SLVS-like der(8) but with a concomitant 8p22p23.1 duplication

De novo dir dup/del of 18q characterized by SNP arrays and FISH in a girl child with mixed phenotypes

1Laboratorio de Citogenomica y Microarreglos, Departamento de Bioquimica y Medicina Molecular, Facultad de Medicina, Universidad Autonoma de Nuevo Leon, Monterrey, Nuevo Leon 64460, Mexico 2Unidad de Biologia Molecular, Genomica y Secuenciacion, Centro de Investigacion y Desarrollo en Ciencias de la Salud, Universidad Autonoma de Nuevo Leon, Monterrey, Nuevo Leon 64460, Mexico 3Laboratorio de Genetica, Hospital General de Culiacan, SSS, Culiacan 80019, Sinaloa, Mexico 4Centro de Rehabilitacion y Educacion Especial (CREE), DIF, Sinaloa 80019, Mexico 5Division de Genetica, Instituto Mexicano del Seguro Social, CIBO, Guadalajara 44340, Mexico 6Doctorado en Genetica Humana, CUCS, Universidad de Guadalajara, Guadalajara 44340, Mexico 7Laboratorio de Genetica y Biologia Molecular, FCQB, Universidad Autonoma de Sinaloa, Sinaloa 80019, Mexico 8Laboratorio de Genetica Humana, Unidad de Investigacion, Facultad de Medicina, Universidad Autonoma de Sinaloa, Sinaloa 80019, Mexico