Sandrine Pereira

Combined treatment with statins and aminobisphosphonates extends longevity in a mouse model of human premature aging

Several human progerias, including Hutchinson-Gilford progeria syndrome (HGPS), are caused by the accumulation at the nuclear envelope of farnesylated forms of truncated prelamin A, a protein that is also altered during normal aging. Previous studies in cells from individuals with HGPS have shown that farnesyltransferase inhibitors (FTIs) improve nuclear abnormalities associated with prelamin A accumulation, suggesting that these compounds could represent a therapeutic approach for this devastating progeroid syndrome. We show herein that both prelamin A and its truncated form progerin/LAΔ50 undergo alternative prenylation by geranylgeranyltransferase in the setting of farnesyltransferase inhibition, which could explain the low efficiency of FTIs in ameliorating the phenotypes of progeroid mouse models. We also show that a combination of statins and aminobisphosphonates efficiently inhibits both farnesylation and geranylgeranylation of progerin and prelamin A and markedly improves the aging-like phenotypes of mice deficient in the metalloproteinase Zmpste24, including growth retardation, loss of weight, lipodystrophy, hair loss and bone defects. Likewise, the longevity of these mice is substantially extended. These findings open a new therapeutic approach for human progeroid syndromes associated with nuclear-envelope abnormalities.

Protein kinase CK2 contributes to the organization of sodium channels in axonal membranes by regulating their interactions with ankyrin G

In neurons, generation and propagation of action potentials requires the precise accumulation of sodium channels at the axonal initial segment (AIS) and in the nodes of Ranvier through ankyrin G scaffolding. We found that the ankyrin-binding motif of Nav1.2 that determines channel concentration at the AIS depends on a glutamate residue (E1111), but also on several serine residues (S1112, S1124, and S1126). We showed that phosphorylation of these residues by protein kinase CK2 (CK2) regulates Nav channel interaction with ankyrins. Furthermore, we observed that CK2 is highly enriched at the AIS and the nodes of Ranvier in vivo. An ion channel chimera containing the Nav1.2 ankyrin-binding motif perturbed endogenous sodium channel accumulation at the AIS, whereas phosphorylation-deficient chimeras did not. Finally, inhibition of CK2 activity reduced sodium channel accumulation at the AIS of neurons. In conclusion, CK2 contributes to sodium channel organization by regulating their interaction with ankyrin G.

SHAPE AND TEXTURE INDEXES APPLICATION TO CELL NUCLEI CLASSIFICATION

This paper describes the sequence of construction of a cell nuclei classification model by the analysis, the characterization and the classification of shape and texture. We describe first the elaboration of dedicated shape indexes and second the construction of the associated classification submodel. Then we present a new method of texture characterization, based on the construction and the analysis of statistical matrices encoding the texture. The various characterization techniques developed in this paper are systematically compared to previous approaches. In particular, we paid special attention to the results obtained by a versatile classification method using a large range of descriptors dedicated to the characterization of shapes and textures. Finally, the last classifier built with our methods achieved 88% of classification out of the 94% possible.

Influence of Nucleoshuttling of the ATM Protein in the Healthy Tissues Response to Radiation Therapy: Toward a Molecular Classification of Human Radiosensitivity

PURPOSE Whereas post-radiation therapy overreactions (OR) represent a clinical and societal issue, there is still no consensual radiobiological endpoint to predict clinical radiosensitivity. Since 2003, skin biopsy specimens have been collected from patients treated by radiation therapy against different tumor localizations and showing a wide range of OR. Here, we aimed to establish quantitative links between radiobiological factors and OR severity grades that would be relevant to radioresistant and genetic hyperradiosensitive cases. METHODS AND MATERIALS Immunofluorescence experiments were performed on a collection of skin fibroblasts from 12 radioresistant, 5 hyperradiosensitive, and 100 OR patients irradiated at 2 Gy. The numbers of micronuclei, γH2AX, and pATM foci that reflect different steps of DNA double-strand breaks (DSB) recognition and repair were assessed from 10 minutes to 24 hours after irradiation and plotted against the severity grades established by the Common Terminology Criteria for Adverse Events and the Radiation Therapy Oncology Group. RESULTS OR patients did not necessarily show a gross DSB repair defect but a systematic delay in the nucleoshuttling of the ATM protein required for complete DSB recognition. Among the radiobiological factors, the maximal number of pATM foci provided the best discrimination among OR patients and a significant correlation with each OR severity grade, independently of tumor localization and of the early or late nature of reactions. CONCLUSIONS Our results are consistent with a general classification of human radiosensitivity based on 3 groups: radioresistance (group I); moderate radiosensitivity caused by delay of nucleoshuttling of ATM, which includes OR patients (group II); and hyperradiosensitivity caused by a gross DSB repair defect, which includes fatal cases (group III).

Low Doses of Gamma-Irradiation Induce an Early Bystander Effect in Zebrafish Cells Which Is Sufficient to Radioprotect Cells

The term “bystander effect” is used to describe an effect in which cells that have not been exposed to radiation are affected by irradiated cells though various intracellular signaling mechanisms. In this study we analyzed the kinetics and mechanisms of bystander effect and radioadaptation in embryonic zebrafish cells (ZF4) exposed to chronic low dose of gamma rays. ZF4 cells were irradiated for 4 hours with total doses of gamma irradiation ranging from 0.01–0.1 Gy. In two experimental conditions, the transfer of irradiated cells or culture medium from irradiated cells results in the occurrence of DNA double strand breaks in non-irradiated cells (assessed by the number of γ-H2AX foci) that are repaired at 24 hours post-irradiation whatever the dose. At low total irradiation doses the bystander effect observed does not affect DNA repair mechanisms in targeted and bystander cells. An increase in global methylation of ZF4 cells was observed in irradiated cells and bystander cells compared to control cells. We observed that pre-irradiated cells which are then irradiated for a second time with the same doses contained significantly less γ-H2AX foci than in 24 h gamma-irradiated control cells. We also showed that bystander cells that have been in contact with the pre-irradiated cells and then irradiated alone present less γ-H2AX foci compared to the control cells. This radioadaptation effect is significantly more pronounced at the highest doses. To determine the factors involved in the early events of the bystander effect, we performed an extensive comparative proteomic study of the ZF4 secretomes upon irradiation. In the experimental conditions assayed here, we showed that the early events of bystander effect are probably not due to the secretion of specific proteins neither the oxidation of these secreted proteins. These results suggest that early bystander effect may be due probably to a combination of multiple factors.

Clinical and genetic analysis of a new multigenerational pedigree with GEFS+ (Generalized Epilepsy with Febrile Seizures Plus).

Summary: Febrile seizures affect 2–5% of all children younger than 6 years. A small proportion of children with febrile seizures later develop epilepsy. The syndrome of generalized epilepsy with febrile seizures plus (GEFS+) is a heterogeneous disorder characterized by febrile seizures that may persist beyond age 6 years and nonfebrile seizures. Several genes have been localized for FS by linkage analysis, and three GEFS+ genes (SCN1A, SCN1B, GABRG2) have been identified. We identified a large multigenerational family with GEFS+ in France. All affected members had FSs. Among them, seven had other types of epileptic seizures including FSs after age 6 years, nonfebrile generalized seizures, or partial seizures later in life. Genetic linkage study excluded the candidate genes and loci for FS and GEFS+, thus proving the existence of a new GEFS+ genetic locus underlying the phenotype observed in this family.

Complete Loss of the Cytoplasmic Carboxyl Terminus of the KCNQ2 Potassium Channel: A Novel Mutation in a Large Czech Pedigree with Benign Neonatal Convulsions or Other Epileptic Phenotypes

Summary:  Purpose: Benign neonatal familial convulsions (BNFCs) represent a rare epileptic disorder with autosomal dominant mode of inheritance. To date, two voltage‐gated potassium (K+) channel genes, KCNQ2 and KCNQ3, have been identified in typical BNFC families. The study of new pedigrees may help detect new mutations and define genotype–phenotype correlations.

Nuclear localization of a novel human syntaxin 1B isoform.

The syntaxins are proteins associated with various intracellular membrane compartments. They are major participants in a large variety of physiological processes where membrane fusion occurs, including exocytosis. We have identified a novel syntaxin isoform generated by alternative splicing of the human STX1B gene. In contrast with the canonical syntaxins, this isoform (STX1B-DeltaTMD) lacked the classical C-terminal transmembrane domain and localized to the nucleus of various tumoral and non-tumoral cell types including human brain cortical neurons in vivo. The reversible blockade of STX1B-DeltaTMD nuclear import demonstrated that nuclear import occurred via a Ran-dependent pathway. A specific and glycine-rich C-terminus of 15 amino acids served as an unconventional nuclear localization signal. STX1B-DeltaTMD colocalized with Lamin A/C and NuMA (NUclear Mitotic Apparatus protein) in interphasic nuclei, and with NuMA and gamma-tubulin in the pericentrosomal region of the mitotic spindle in dividing cells. In a series of 37 human primary brain tumors, the ratio of STX1B-DeltaTMD to Lamin A/C transcripts was a significant prognostic marker of survival, independent of tumor staging. The characterization of STX1B-DeltaTMD as the first nucleoplasmic syntaxin with no transmembrane domain, illustrates the importance of alternative splicing in the emergence of unsuspected properties of the syntaxins in human cells, in both physiological and pathological conditions.

Effects of chronic exposure to environmentally relevant concentrations of waterborne depleted uranium on the digestive tract of zebrafish, Danio rerio

Uranium is a naturally occurring element, but activities linked to the nuclear fuel cycle can increase background levels in the surrounding waters. For this reason it is important to understand how this affects organisms residing in the water column. The objective of this study was to assess histopathological effects of uranium on the gut wall of a widely used model organism: zebrafish, Danio rerio. To this end we exposed zebrafish to 84 and 420 nM depleted uranium for over a month and then examined the histology of intestines of exposed individuals compared to controls. The gut wall of individuals exposed to 84 and 420 nM of uranium had large regions of degraded mucosa. Using transmission electron microscopy (TEM) coupled to energy-dispersive X-ray spectroscopy microanalysis (EDX) we found that uranium induced a decrease in the amount of calcium containing mitochondrial matrix granules per mitochondria. This is suggestive of perturbations to cellular metabolism and more specifically to cellular calcium homeostasis. TEM-EDX of the gut wall tissue further showed that some uranium was internalized in the nucleus of epithelial cells in the 420 nM treatment. Fluorescent in situ hybridization using specific probes to detect all eubacteria was performed on frozen sections of 6 individual fish in the 84 nM and 420 nM treatments. Bacterial colonization of the gut of individuals in the 420 nM seemed to differ from that of the controls and 84 nM individuals. We suggest that host-microbiota interactions are potentially disturbed in response to uranium induced stress. The damage induced by waterborne uranium to the gut wall did not seem to depend on the concentration of uranium in the media. We measure whole body residues of uranium at the end of the experiment and compute the mean dose rate absorbed for each condition. We discuss why effects might be uncoupled from external concentration and highlight that it is not so much the external concentration but the dynamics of internalization which are important players in the game.

Epilepsy and deletions at chromosome 2q24

Chromosomal abnormalities are an important cause of epilepsy [Singh et al., 2002], which might be the presenting symptom in the context of a specific syndrome. A recent article by Langer et al. [2006] reports on a translocation t(2;15) with deletion at 2q24-q31 in a girl with epilepsy, dysmorphic features, and severe developmental delay. As this case adds to the growing list of severe epilepsy associated with deletions at chromosome 2q, it deserves complementary information and comments. We would like to point out that wedescribed a very similar case in a previous report on a girl carrying a deletion at chromosome 2q24 [Pereira et al., 2004]. In our patient as well as in the patient described by Langer et al. [2006], epileptic seizures first started early in life, at 2 and 3 months 1/2, respectively. Seizures were frequent in both cases and were accompanied by severe apnea. Both children had severe developmental delay and no speech development. In the two patients, additional common features were encountered, including microcephaly, downslanting and small palpebral fissures, and abnormal ears. Despite the clear similarities existing between those two cases, other clinical manifestations were not shared in common by the two patients. Our patient had single palmar creases bilaterally and partial syndactyly between the 2nd and 3rd toes, and cardiac examination showed a small interventricular communication. In contrast, the patient in Langer et al. [2006] had micrognathia, blepharophimosis, hypertelorism, and coloboma of the iris and retina. This is reminiscent of the clinical features described in other cases of del(2)(q31q33) [Ramer et al., 1990] and del(2)(q24–q31) [Nixon et al., 1997]. The size and the boundaries of the respective deleted areas vary from one patient to another (Fig. 1) and this might well explain the clinical differences described above. In the case of the patient described by Langer et al. [2006], the translocation breakpoint at 15q14 might also participate in the phenotype. In this latter patient, the deleted region on chromosome 2 spans about 13 Mb between markers rs198688 and rs1399959 and contains up to 76 genes including SCN1A and SCN2A, the mutations of which have already been associated with various epileptic syndromes [Meisler and Kearney, 2005]. Our critical region was of smaller size and spanned3–7Mb [Pereira et al., 2004] (Fig. 1). It is fully included within the 13 Mb-deleted area described by Langer et al. [2006] (Fig. 1) and ‘only’ contains 32 genes at most and 10 genes at least, including SCN1A and SCN2A as well as three other sodium channel genes (see the human genome sequence at UCSC web site: http://genome.ucsc. edu) [Pereira et al., 2004]. Epilepsy was not seen in the patient reported by Nixon et al. [1997]. The data reported by Langer et al. [2006] thus reinforce our previous hypothesis [Pereira et al., 2004], that the epileptic phenotype in these patients is specifically associated with the deletion of the genomic area comprised between marker D2S354 (marker d in Fig. 1) and marker D2S2345 (h) at most. This genomic region actually comprises the cluster of five sodium channel genes mentioned above. As already discussed [Pereira et al., 2004; Langer et al., 2006], the seizures in the affected children might well be due to loss or haploinsufficiency of one or several of these genes. Such deletion events might remain undetected and hence might be