PubMed Health⌕ Search

Biomedical subjects

Annie Andrieux

Publications and source records attributed to Annie Andrieux.

15 recordsLinked to original sources
PubMed2006

Post-translational arginylation of calreticulin: a new isospecies of calreticulin component of stress granules.

Post-translational arginylation consists of the covalent union of an arginine residue to a Glu, Asp, or Cys amino acid at the N-terminal position of proteins. This reaction is catalyzed by the enzyme arginyl-tRNA protein transferase. Using mass spectrometry, we have recently demonstrated in vitro the post-translational incorporation of arginine into the calcium-binding protein calreticulin (CRT). To further study arginylated CRT we raised an antibody against the peptide (RDPAIYFK) that contains an arginine followed by the first 7 N-terminal amino acids of mature rat CRT. This antibody specifically recognizes CRT obtained from rat soluble fraction that was arginylated in vitro and also recognizes endogenous arginylated CRT from NIH 3T3 cells in culture, indicating that CRT arginylation takes place in living cells. Using this antibody we found that arginylation of CRT is Ca2+-regulated. In vitro and in NIH 3T3 cells in culture, the level of arginylated CRT increased with the addition of a Ca2+ chelator to the medium, whereas a decreased arginine incorporation into CRT was found in the presence of Ca2+. The arginylated CRT was observed in the cytosol, in contrast to the non-arginylated CRT that is in the endoplasmic reticulum. Under stress conditions, arginylated CRT was found associated to stress granules. These results suggest that CRT arginylation occurs in the cytosolic pool of mature CRT (defined by an Asp acid N-terminal) that is probably retrotranslocated from the endoplasmic reticulum.

Animals↗
PubMed2006

Altered expression of synaptic protein mRNAs in STOP (MAP6) mutant mice.

Stable tubule-only polypeptide (STOP) proteins are a family of microtubule associated proteins (MAPs) important in microtubule stabilization. Data indicating a role for microtubules in synaptic function has come from studies of the STOP null mouse, which exhibits synaptic deficits, in association with behavioural changes that are alleviated by antipsychotic treatment. These findings suggested that STOP mutant mice may be useful in studies of synaptic function, and could be especially relevant to schizophrenia, postulated to be a disorder of the synapse. Moreover, a genetic association between STOP and schizophrenia has been reported. This study aimed to further characterize synaptic alterations in STOP null and heterozygous mice. Using in situ hybridization histochemistry, the mRNA expression of three pre-synaptic (synaptophysin; growth associated protein-43 (GAP-43); vesicular glutamate transporter-1 (VGlut1)) and two post-synaptic (spinophilin; MAP2) proteins, was quantified in female STOP null (n = 7), heterozygous (n = 5) and wild type (n = 6) mice. For STOP null and heterozygous mice, synaptophysin, VGlut1, GAP-43 and spinophilin mRNAs were decreased in the hippocampus, whilst in addition in the null mice, synaptophysin, VGlut1 and spinophilin mRNAs were decreased in the cerebellum. Alterations in synaptic protein mRNA expression were also detected in the frontal and occipital cortex. MAP2 mRNA expression was unchanged in all brain regions. The profile of mRNA changes is broadly similar to that observed in schizophrenia. Together the data provide supporting evidence for a role for microtubules in synaptic function, and suggest that STOP, or other microtubule proteins, may contribute to the synaptic pathology of schizophrenia.

Animals↗
PubMed2006

Tubulin tyrosination is a major factor affecting the recruitment of CAP-Gly proteins at microtubule plus ends.

Tubulin-tyrosine ligase (TTL), the enzyme that catalyzes the addition of a C-terminal tyrosine residue to alpha-tubulin in the tubulin tyrosination cycle, is involved in tumor progression and has a vital role in neuronal organization. We show that in mammalian fibroblasts, cytoplasmic linker protein (CLIP) 170 and other microtubule plus-end tracking proteins comprising a cytoskeleton-associated protein glycine-rich (CAP-Gly) microtubule binding domain such as CLIP-115 and p150 Glued, localize to the ends of tyrosinated microtubules but not to the ends of detyrosinated microtubules. In vitro, the head domains of CLIP-170 and of p150 Glued bind more efficiently to tyrosinated microtubules than to detyrosinated polymers. In TTL-null fibroblasts, tubulin detyrosination and CAP-Gly protein mislocalization correlate with defects in both spindle positioning during mitosis and cell morphology during interphase. These results indicate that tubulin tyrosination regulates microtubule interactions with CAP-Gly microtubule plus-end tracking proteins and provide explanations for the involvement of TTL in tumor progression and in neuronal organization.

Animals↗
PubMed2006

STOP-like protein 21 is a novel member of the STOP family, revealing a Golgi localization of STOP proteins.

Neuronal microtubules are stabilized by two calmodulin-regulated microtubule-associated proteins, E-STOP and N-STOP, which when suppressed in mice induce severe synaptic and behavioral deficits. Here we show that mature neurons also contain a 21-kDa STOP-like protein, SL21, which shares calmodulin-binding and microtubule-stabilizing homology domains with STOP proteins. Accordingly, in different biochemical or cellular assays, SL21 has calmodulin binding and microtubule stabilizing activity. However, in cultured hippocampal neurons, SL21 antibodies principally stain the somatic Golgi and punctate Golgi material in neurites. In cycling cells, transfected SL21 decorates microtubules when expressed at high levels but is otherwise principally visible at the Golgi. The Golgi targeting of SL21 depends on the presence of cysteine residues located within the SL21 N-terminal domain, suggesting that Golgi targeting may require SL21 palmitoylation. Accordingly we find that SL21 is palmitoylated in vivo. N-STOP and E-STOP, which contain the Golgi targeting sequences present in SL21, also display distinct Golgi staining when expressed at low level in cycling cells. Thus neuronal proteins of the STOP family have the capacity to associate with Golgi material, which could be important for STOP synaptic functions.

Amino Acid Sequence↗
PubMed2006

Microtubule stabilizer ameliorates synaptic function and behavior in a mouse model for schizophrenia.

BACKGROUND: Recent data suggest that cytoskeletal defects may play a role in schizophrenia. We previously imitated features of schizophrenia in an animal model by disrupting gene coding for a microtubule-associated protein called STOP. STOP-null mice display synaptic defects in glutamatergic neurons, hyper-dopaminergy, and severe behavioral disorders. Synaptic and behavioral deficits are amended by neuroleptic treatment in STOP-null mice, providing an attractive model to test new antipsychotic agents. We examined the effects of a taxol-related microtubule stabilizer, epothilone D. METHODS: Mice were treated either with vehicle alone or with epothilone D. Treatment effects on synaptic function were assessed using electron-microscopy quantification of synaptic vesicle pools and electrophysiology in the CA1 region of the hippocampus. Dopamine transmission was investigated using electrochemical assays. Behavior was principally assessed using tests of maternal skills. RESULTS: In STOP-null mice, treatment with epothilone D increased synaptic vesicle pools, ameliorated both short- and long-term forms of synaptic plasticity in glutamatergic neurons, and had a dramatic beneficial effect on mouse behavior. CONCLUSIONS: A microtubule stabilizer can have a beneficial effect on synaptic function and behavior, suggesting new possibilities for treatment of schizophrenia.

Animals↗
PubMed2006

Phosphorylation of microtubule-associated protein STOP by calmodulin kinase II.

STOP proteins are microtubule-associated, calmodulin-regulated proteins responsible for the high degree of stabilization displayed by neuronal microtubules. STOP suppression in mice induces synaptic defects affecting both short and long term synaptic plasticity in hippocampal neurons. Interestingly, STOP has been identified as a component of synaptic structures in neurons, despite the absence of microtubules in nerve terminals, indicating the existence of mechanisms able to induce a translocation of STOP from microtubules to synaptic compartments. Here we have tested STOP phosphorylation as a candidate mechanism for STOP relocalization. We show that, both in vitro and in vivo, STOP is phosphorylated by the multifunctional enzyme calcium/calmodulin-dependent protein kinase II (CaMKII), which is a key enzyme for synaptic plasticity. This phosphorylation occurs on at least two independent sites. Phosphorylated forms of STOP do not bind microtubules in vitro and do not co-localize with microtubules in cultured differentiating neurons. Instead, phosphorylated STOP co-localizes with actin assemblies along neurites or at branching points. Correlatively, we find that STOP binds to actin in vitro. Finally, in differentiated neurons, phosphorylated STOP co-localizes with clusters of synaptic proteins, whereas unphosphorylated STOP does not. Thus, STOP phosphorylation by CaMKII may promote STOP translocation from microtubules to synaptic compartments where it may interact with actin, which could be important for STOP function in synaptic plasticity.

Actins↗
PubMed2005

STOP knockout and NMDA NR1 hypomorphic mice exhibit deficits in sensorimotor gating.

Schizophrenia is a chronic and debilitating disease which is thought to arise from a neuro-developmental disorder. Both the stable tubule-only polypeptide (STOP) protein and the N-methyl-D-aspartate (NMDA) NR1 subunit are involved in neuronal development and physiology. It has therefore been postulated that transgenic mice lacking either the STOP or the NMDAR1 gene would show a 'schizophrenic-like' phenotype. Here, STOP knockout and NMDA NR1 hypomorphic mice were assessed in a behavioural measure that can be used to detect schizophrenic-like phenotypes: a change in sensorimotor gating, measured through prepulse inhibition (PPI). STOP knockout mice were further assessed in another measure of 'schizophrenic-like behaviour': hyperlocomotion. The PPI deficit exhibited by both the STOP knockout and NMDA knockdown mice could not be reversed by acute treatment with the atyptical antipsychotic, clozapine (1 mg/kg, i.p.) but the hyperlocomotion shown by the STOP knockout mice was reversed with the same acute dose of clozapine.

Acoustic Stimulation↗
PubMed2005

A vital role of tubulin-tyrosine-ligase for neuronal organization.

Tubulin is subject to a special cycle of detyrosination/tyrosination in which the C-terminal tyrosine of alpha-tubulin is cyclically removed by a carboxypeptidase and readded by a tubulin-tyrosine-ligase (TTL). This tyrosination cycle is conserved in evolution, yet its physiological importance is unknown. Here, we find that TTL suppression in mice causes perinatal death. A minor pool of tyrosinated (Tyr-)tubulin persists in TTL null tissues, being present mainly in dividing TTL null cells where it originates from tubulin synthesis, but it is lacking in postmitotic TTL null cells such as neurons, which is apparently deleterious because early death in TTL null mice is, at least in part, accounted for by a disorganization of neuronal networks, including a disruption of the cortico-thalamic loop. Correlatively, cultured TTL null neurons display morphogenetic anomalies including an accelerated and erratic time course of neurite outgrowth and a premature axonal differentiation. These anomalies may involve a mislocalization of CLIP170, which we find lacking in neurite extensions and growth cones of TTL null neurons. Our results demonstrate a vital role of TTL for neuronal organization and suggest a requirement of Tyr-tubulin for proper control of neurite extensions.

Animals↗
PubMed2005

MCAK associates with the tips of polymerizing microtubules.

MCAK is a member of the kinesin-13 family of microtubule (MT)-depolymerizing kinesins. We show that the potent MT depolymerizer MCAK tracks (treadmills) with the tips of polymerizing MTs in living cells. Tip tracking of MCAK is inhibited by phosphorylation and is dependent on the extreme COOH-terminal tail of MCAK. Tip tracking is not essential for MCAK's MT-depolymerizing activity. We propose that tip tracking is a mechanism by which MCAK is preferentially localized to regions of the cell that modulate the plus ends of MTs.

Animals↗
PubMed2005

Dopaminergic transmission in STOP null mice.

Neuroleptics are thought to exert their anti-psychotic effects by counteracting a hyper-dopaminergic transmission. Here, we have examined the dopaminergic status of STOP (stable tubule only polypeptide) null mice, which lack a microtubule-stabilizing protein and which display neuroleptic-sensitive behavioural disorders. Dopamine transmission was investigated using both behavioural analysis and measurements of dopamine efflux in different conditions. Compared to wild-type mice in basal conditions or following mild stress, STOP null mice showed a hyper-locomotor activity, which was erased by neuroleptic treatment, and an increased locomotor reactivity to amphetamine. Such a behavioural profile is indicative of an increased dopaminergic transmission. In STOP null mice, the basal dopamine concentrations, measured by quantitative microdialysis, were normal in both the nucleus accumbens and the striatum. When measured by electrochemical techniques, the dopamine efflux evoked by electrical stimulations mimicking physiological stimuli was dramatically increased in the nucleus accumbens of STOP null mice, apparently due to an increased dopamine release, whereas dopaminergic uptake and auto-inhibition mechanisms were normal. In contrast, dopamine effluxes were slightly diminished in the striatum. Together with previous results, the present study indicates the association in STOP null mice of hippocampal hypo-glutamatergy and of limbic hyper-dopaminergy. Such neurotransmission defects are thought to be central to mental diseases such as schizophrenia.

Action Potentials↗
PubMed2003

STOP proteins.

Microtubules assembled from purified tubulin in vitro are labile, rapidly disassembling when exposed to a variety of depolymerizing conditions such as cold temperature. In contrast, in many cell types, microtubules seem to be unaffected when the cell is exposed to the cold. This resistance of microtubules to the cold has been intriguing because the earliest and by far most studied microtubule-associated proteins such as MAP2 and tau are devoid of microtubule cold stabilizing activity. Over the past several years, it has been shown that resistance of microtubules to the cold is largely due to polymer association with a class of microtubule-associated proteins called STOPs. STOPs are calmodulin-binding and calmodulin-regulated proteins which, in mammals, are encoded by a single gene but exhibit substantial cell specific variability due to mRNA splicing and alternative promoter use. STOP microtubule stabilizing activity has been ascribed to two classes of new bifunctional calmodulin- and microtubule-binding motifs, with distinct microtubule binding properties in vivo. STOPs seem to be restricted to vertebrates and are composed of a conserved domain split by the apparent insertion of variable sequences that are completely unrelated among species. Recently, STOP suppression in mice has been found to induce synaptic defects associated with neuroleptic-sensitive behavioral disorders. Thus, STOPs are important for synaptic plasticity. Additionally, STOP-deficient mice may yield a pertinent model for the study of neuroleptics in illnesses such as schizophrenia, currently thought to result from defects in synapse function.

Amino Acid Sequence↗
PubMed2003

CD38-dependent ADP-ribosyl cyclase activity in developing and adult mouse brain.

CD38 is a transmembrane glycoprotein that is expressed in many tissues throughout the body. In addition to its major NAD+-glycohydrolase activity, CD38 is also able to synthesize cyclic ADP-ribose, an endogenous calcium-regulating molecule, from NAD+. In the present study, we have compared ADP-ribosyl cyclase and NAD+-glycohydrolase activities in protein extracts of brains from developing and adult wild-type and Cd38 -/- mice. In extracts from wild-type brain, cyclase activity was detected spectrofluorimetrically, using nicotinamide-guanine dinucleotide as a substrate (GDP-ribosyl cyclase activity), as early as embryonic day 15. The level of cyclase activity was similar in the neonate brain (postnatal day 1) and then increased greatly in the adult brain. Using [14C]NAD+ as a substrate and HPLC analysis, we found that ADP-ribose is the major product formed in the brain at all developmental stages. Under the same experimental conditions, neither NAD+-glycohydrolase nor GDP-ribosyl cyclase activity could be detected in extracts of brains from developing or adult Cd38 -/- mice, demonstrating that CD38 is the predominant constitutive enzyme endowed with these activities in brain at all developmental stages. The activity measurements correlated with the level of CD38 transcripts present in the brains of developing and adult wild-type mice. Using confocal microscopy we showed, in primary cultures of hippocampal cells, that CD38 is expressed by both neurons and glial cells, and is enriched in neuronal perikarya. Intracellular NAD+-glycohydrolase activity was measured in hippocampal cell cultures, and CD38-dependent cyclase activity was higher in brain fractions enriched in intracellular membranes. Taken together, these results lead us to speculate that CD38 might have an intracellular location in neural cells in addition to its plasma membrane location, and may play an important role in intracellular cyclic ADP-ribose-mediated calcium signalling in brain tissue.

ADP-ribosyl Cyclase↗
PubMed2003

Overlap of promoter and coding sequences in the mouse STOP gene (Mtap6).

The microtubule-associated proteins STOP are essential for synaptic plasticity and integrated brain function. The STOP gene (Mtap6) is composed of 4 exons and presents different developmental and tissue specific alternative transcripts resulting in the neuronal isoforms E- and N-STOP, and in the fibroblastic F-STOP isoform. We now characterize the transcription initiation sites for neuronal and non neuronal STOP transcripts. Our results show that there is a single neuronal specific promoter for transcription of E- and N-STOP mRNAs. F-STOP mRNA transcription is regulated by a distinct promoter. A remarkable feature of this promoter is that it lies within coding sequences expressed in neuronal E- and N-STOP isoforms.

Animals↗
PubMed2002

The suppression of brain cold-stable microtubules in mice induces synaptic defects associated with neuroleptic-sensitive behavioral disorders.

Neurons contain abundant subsets of highly stable microtubules that resist depolymerizing conditions such as exposure to the cold. Stable microtubules are thought to be essential for neuronal development, maintenance, and function. Previous work has indicated an important role of the microtubule-associated protein STOP in the induction of microtubule cold stability. Here, we developed STOP null mice. These mice were devoid of cold-stable microtubules. In contrast to our expectations, STOP-/- mice had no detectable defects in brain anatomy but showed synaptic defects, with depleted synaptic vesicle pools and impaired synaptic plasticity, associated with severe behavioral disorders. A survey of the effects of psychotropic drugs on STOP-/- mice behavior showed a remarkable and specific effect of long-term administration of neuroleptics in alleviating these disorders. This study demonstrates that STOP is a major factor responsible for the intriguing stability properties of neuronal microtubules and is important for synaptic plasticity. Additionally, STOP-/- mice may yield a pertinent model for study of neuroleptics in illnesses such as schizophrenia, currently thought to result from synaptic defects.

Animals↗