[Comparison of platelet monitoring in patients receiving thromboprophylaxis with fondaparinux or a low molecular weight heparin. The Ariane study]. / Étude Ariane : comparaison de la surveillance plaquettaire chez des patients recevant une thromboprophylaxie par fondaparinux ou HBPM.

OBJECTIVE: To describe and compare the conditions of use of fondaparinux and low molecular weight heparin (LMWH) in the prevention of venous thromboembolism in routine general practice with a focus on platelet monitoring. METHOD: This was an observational and pharmaco-epidemiological survey, performed in France in general practice in adult patients receiving thromboprophylaxis with fondaparinux or a LMWH. The study collected data on medical conditions justifying thromboprophylaxis, reasons for platelet monitoring and type of prescription. RESULTS: Four hundred and seventy general practioners included 837 analysable patients (450 treated with fondaparinux and 387 with LMWH). In the fondaparinux group, the mean age was 61.5±17.3 and 259 (57.6 %) patients were women. In the LMWH group, the mean age was 61.7±17.8 and 205 (53.0 %) patients were women. The reasons of prescribing were: bedridden related to a severe acute medical illness in 255 (56.7 %) patients with fondaparinux and 244 (63.1 %) with LMWH, and reduction of mobility associated with trauma without fracture respectively in 121 (26.9 %) and 85 (22.0 %) of patients. Associated risk factors were varicose veins, obesity and a history of thrombosis. Platelet monitoring was prescribed in 168 (37.6 %) patients treated with fondaparinux. In this group, these prescription were considered "appropriate" in 94 (20.9 %) patients, of whom 76 (16.9 %) were monitored for screening purposes, and "not appropriate" in 67 (14.9 %) patients, because prescribed to monitor thrombo-prophylaxis. In the LMWH group, a platelet count was prescribed in 370 (96.1 %) patients, of whom 312 (81.0 %) receiving a prescription only in order to monitor thromboprophylaxis. DISCUSSION: The results provided in the Ariane study were coherent with literature data (Etape and Depart studies). In comparison with the CNAM study, which evaluated prescription practices for LMWH in thromboprophylaxis in France in 1999, and which reported a global rate of platelet monitoring of 70.0 %, the rate reported in the Ariane study (81.0 %) seems to represent an improvement in the practice standards. Since 2009, Afssaps does not recommend a systematic monitoring with LMWH at acute or prophylactic dose, outside a post surgical context or in case of pre-treatment with unfractionated heparin. CONCLUSION: The Ariane study provides important information on platelet monitoring in patients treated with fondaparinux or LMWH, and also on thromboprohylaxis in general practice.

MTA is an Arabidopsis messenger RNA adenosine methylase and interacts with a homolog of a sex-specific splicing factor.

N6-Methyladenosine is a ubiquitous modification identified in the mRNA of numerous eukaryotes, where it is present within both coding and noncoding regions. However, this base modification does not alter the coding capacity, and its biological significance remains unclear. We show that Arabidopsis thaliana mRNA contains N6-methyladenosine at levels similar to those previously reported for animal cells. We further show that inactivation of the Arabidopsis ortholog of the yeast and human mRNA adenosine methylase (MTA) results in failure of the developing embryo to progress past the globular stage. We also demonstrate that the arrested seeds are deficient in mRNAs containing N6-methyladenosine. Expression of MTA is strongly associated with dividing tissues, particularly reproductive organs, shoot meristems, and emerging lateral roots. Finally, we show that MTA interacts in vitro and in vivo with At FIP37, a homolog of the Drosophila protein FEMALE LETHAL2D and of human WILMS' TUMOUR1-ASSOCIATING PROTEIN. The results reported here provide direct evidence for an essential function for N6-methyladenosine in a multicellular eukaryote, and the interaction with At FIP37 suggests possible RNA processing events that might be regulated or altered by this base modification.

ATM regulates the length of individual telomere tracts in Arabidopsis.

Telomeres have the paradoxical ability of protecting linear chromosome ends from DNA damage sensors by using these same proteins as essential components of their maintenance machinery. We have previously shown that the absence of ataxia telangiectasia mutated (ATM), a central regulator of the DNA damage response, accelerates the onset of genome instability in telomerase-deficient Arabidopsis, without increasing the rate of bulk telomere shortening. Here, we examine individual telomere tracts through successive plant generations using both fluorescence situ in hybridization (FISH) and primer extension telomere repeat amplification (PETRA). Unexpectedly, we found that the onset of profound developmental defects and abundant end-to-end chromosome fusions in fifth generation (G(5)) atm tert mutants required the presence of only one critically shortened telomere. Parent progeny analysis revealed that the short telomere arose as a consequence of an unusually large telomere rapid deletion (TRD) event. The most dramatic TRD was detected in atm tert mutants that had undergone meiosis. Notably, in contrast to TRD, alternative lengthening of telomeres (ALT) was suppressed in the absence of ATM. Finally, we show that size differences between telomeres on homologous chromosome ends are greater for atm tert than tert plants. Altogether, these findings suggest a dual role for ATM in regulating telomere size by promoting elongation of short telomeres and by preventing the accumulation of cells that harbor large telomere deletions.

Arabidopsis POT1 associates with the telomerase RNP and is required for telomere maintenance.

POT1 is a single-copy gene in yeast and humans that encodes a single-strand telomere binding protein required for chromosome end protection and telomere length regulation. In contrast, Arabidopsis harbors multiple, divergent POT-like genes that bear signature N-terminal OB-fold motifs, but otherwise share limited sequence similarity. Here, we report that plants null for AtPOT1 show no telomere deprotection phenotype, but rather exhibit progressive loss of telomeric DNA. Genetic analysis indicates that AtPOT1 acts in the same pathway as telomerase. In vitro levels of telomerase activity in pot1 mutants are significantly reduced and are more variable than wild-type. Consistent with this observation, AtPOT1 physically associates with active telomerase particles. Although low levels of AtPOT1 can be detected at telomeres in unsynchronized cells and in cells arrested in G2, AtPOT1 binding is significantly enhanced during S-phase, when telomerase is thought to act at telomeres. Our findings indicate that AtPOT1 is a novel accessory factor for telomerase required for positive telomere length regulation, and they underscore the coordinate and extraordinarily rapid evolution of telomere proteins and the telomerase enzyme.

ATM and ATR make distinct contributions to chromosome end protection and the maintenance of telomeric DNA in Arabidopsis.

Here we examine the function of ATM and ATR at telomeres in Arabidopsis. Although plants lacking ATM or ATR display wild-type telomere length homeostasis, chromosome end protection is compromised in atm atr mutants. Moreover, atm tert Arabidopsis experience an abrupt, early onset of genome instability, arguing that ATM is required for protection of short telomeres. ATR, by contrast, is required for maintenance of telomeric DNA as telomere shortening is dramatically accelerated in atr tert mutants relative to tert plants. Thus, ATM and ATR make essential and distinct contributions to chromosome end protection and telomere maintenance in higher eukaryotes.

A C-terminal Myb extension domain defines a novel family of double-strand telomeric DNA-binding proteins in Arabidopsis.

Little is known about the protein composition of plant telomeres. We queried the Arabidopsis thaliana genome data base in search of genes with similarity to the human telomere proteins hTRF1 and hTRF2. hTRF1/hTRF2 are distinguished by the presence of a single Myb-like domain in their C terminus that is required for telomeric DNA binding in vitro. Twelve Arabidopsis genes fitting this criterion, dubbed TRF-like (TRFL), fell into two distinct gene families. Notably, TRFL family 1 possessed a highly conserved region C-terminal to the Myb domain called Myb-extension (Myb-ext) that is absent in TRFL family 2 and hTRF1/hTRF2. Immunoprecipitation experiments revealed that recombinant proteins from TRFL family 1, but not those from family 2, formed homodimers and heterodimers in vitro. DNA binding studies with isolated C-terminal fragments from TRFL family 1 proteins, but not family 2, showed specific binding to double-stranded plant telomeric DNA in vitro. Removal of the Myb-ext domain from TRFL1, a family 1 member, abolished DNA binding. However, when the Myb-ext domain was introduced into the corresponding region in TRFL3, a family 2 member, telomeric DNA binding was observed. Thus, Myb-ext is required for binding plant telomeric DNA and defines a novel class of proteins in Arabidopsis.

The immunophilin-interacting protein AtFIP37 from Arabidopsis is essential for plant development and is involved in trichome endoreduplication.

The FKBP12 (FK506-binding protein 12 kD) immunophilin interacts with several protein partners in mammals and is a physiological regulator of the cell cycle. In Arabidopsis, only one specific partner of AtFKBP12, namely AtFIP37 (FKBP12 interacting protein 37 kD), has been identified but its function in plant development is not known. We present here the functional analysis of AtFIP37 in Arabidopsis. Knockout mutants of AtFIP37 show an embryo-lethal phenotype that is caused by a strong delay in endosperm development and embryo arrest. AtFIP37 promoter::beta-glucuronidase reporter gene constructs show that the gene is expressed during embryogenesis and throughout plant development, in undifferentiating cells such as meristem or embryonic cells as well as highly differentiating cells such as trichomes. A translational fusion with the enhanced yellow fluorescent protein indicates that AtFIP37 is a nuclear protein localized in multiple subnuclear foci that show a speckled distribution pattern. Overexpression of AtFIP37 in transgenic lines induces the formation of large trichome cells with up to six branches. These large trichomes have a DNA content up to 256C, implying that these cells have undergone extra rounds of endoreduplication. Altogether, these data show that AtFIP37 is critical for life in Arabidopsis and implies a role for AtFIP37 in the regulation of the cell cycle as shown for FKBP12 and TOR (target of rapamycin) in mammals.