Identification of the SPG15 gene, encoding spastizin, as a frequent cause of complicated autosomal-recessive spastic paraplegia, including Kjellin syndrome

Hereditary spastic paraplegias (HSPs) are genetically and phenotypically heterogeneous disorders. Both "uncomplicated" and "complicated" forms have been described with various modes of inheritance. Sixteen loci for autosomal-recessive "complicated" HSP have been mapped. The SPG15 locus was first reported to account for a rare form of spastic paraplegia variably associated with mental impairment, pigmented maculopathy, dysarthria, cerebellar signs, and distal amyotrophy, sometimes designated as Kjellin syndrome. Here, we report the refinement of SPG15 to a 2.64 Mb genetic interval on chromosome 14q23.3-q24.2 and the identification of ZFYVE26, which encodes a zinc-finger protein with a FYVE domain that we named spastizin, as the cause of SPG15. Six different truncating mutations were found to segregate with the disease in eight families with a phenotype that included variable clinical features of Kjellin syndrome. ZFYVE26 mRNA was widely distributed in human tissues, as well as in rat embryos, suggesting a possible role of this gene during embryonic development. In the adult rodent brain, its expression profile closely resembled that of SPG11, another gene responsible for complicated HSP. In cultured cells, spastizin colocalized partially with markers of endoplasmic reticulum and endosomes, suggesting a role in intracellular trafficking.     

mTORC1 stimulates cell growth through SAM synthesis and m6A mRNA-dependent control of protein synthesis

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A Chemically Defined Medium for Rabbit Embryo Cryopreservation

This study evaluates a new synthetic substitute (CRYO3, Ref. 5617, Stem Alpha, France) for animal-based products in rabbit embryo cryopreservation solutions. This evaluation was performed using two approaches: a thermodynamic approach using differential scanning calorimetry and a biological approach using rabbit embryo slow-freezing. During the experiment, foetal calf serum (FCS) was used as a reference. Because FCS varies widely by supplier, three different FCS were selected for the thermodynamic approach. The rabbit embryo slow-freezing solutions were made from Dulbecco''s phosphate buffer saline containing 1.5 M Dimethyl Sulfoxide and 18% (v.v⁻¹) of CRYO3 or 18% (v.v⁻¹) of FCS. These solutions were evaluated using four characteristics: the end of melting temperature, the enthalpy of crystallisation (thermodynamic approach) and the embryo survival rates after culture and embryo transfer (biological approach). In the thermodynamic approach, the solutions containing one of the three different FCS had similar mean thermodynamic characteristics but had different variabilities in the overall data with aberrant values. The solution containing CRYO3 had similar thermodynamic properties when compared to those containing FCS. Moreover, no aberrant value was measured in the solution containing CRYO3. This solution appears to be more stable than the solutions containing a FCS. In the biological approach, the in vitro embryo survival rates obtained with the solution containing CRYO3 (73.7% and 81.3%) and with the solution containing a FCS (77.6% and 71.9%) were similar (p = 0.7). Nevertheless, during the in vivo evaluation, the implantation rate (21.8%) and the live-foetuses rate (18.8%) of the CRYO3 group were significantly higher than the implantation rate (7.1%, p = 0.0002) and the live-foetuses rate (5.3%, p = 0.0002) of the FCS group. The pregnancy rate was also higher in the CRYO3 group compared to the FCS group (81.3% and 43.8%, respectively, p = 0.066). We conclude that CRYO3 can be used as a chemically defined substitute for animal-based products in rabbit embryo cryopreservation solutions.     

Orthosteric Binding of ρ-Da1a,a Natural Peptide of Snake Venom Interacting Selectively with the α1A-Adrenoceptor

ρ-Da1a is a three-finger fold toxin from green mamba venom that is highly selective for the α_1A-adrenoceptor. This toxin has atypical pharmacological properties, including incomplete inhibition of ³H-prazosin or ¹²⁵I-HEAT binding and insurmountable antagonist action. We aimed to clarify its mode of action at the α_1A-adrenoceptor. The affinity (pKi 9.26) and selectivity of ρ-Da1a for the α_1A-adrenoceptor were confirmed by comparing binding to human adrenoceptors expressed in eukaryotic cells. Equilibrium and kinetic binding experiments were used to demonstrate that ρ-Da1a, prazosin and HEAT compete at the α_1A-adrenoceptor. ρ-Da1a did not affect the dissociation kinetics of ³H-prazosin or ¹²⁵I-HEAT, and the IC₅₀ of ρ-Da1a, determined by competition experiments, increased linearly with the concentration of radioligands used, while the residual binding by ρ-Da1a remained stable. The effect of ρ-Da1a on agonist-stimulated Ca²⁺ release was insurmountable in the presence of phenethylamine- or imidazoline-type agonists. Ten mutations in the orthosteric binding pocket of the α_1A-adrenoceptor were evaluated for alterations in ρ-Da1a affinity. The D106^3.32A and the S188^5.42A/S192^5.46A receptor mutations reduced toxin affinity moderately (6 and 7.6 times, respectively), while the F86^2.64A, F288^6.51A and F312^7.39A mutations diminished it dramatically by 18- to 93-fold. In addition, residue F86^2.64 was identified as a key interaction point for ¹²⁵I-HEAT, as the variant F86^2.64A induced a 23-fold reduction in HEAT affinity. Unlike the M1 muscarinic acetylcholine receptor toxin MT7, ρ-Da1a interacts with the human α_1A-adrenoceptor orthosteric pocket and shares receptor interaction points with antagonist (F86^2.64, F288^6.51 and F312^7.39) and agonist (F288^6.51 and F312^7.39) ligands. Its selectivity for the α_1A-adrenoceptor may result, at least partly, from its interaction with the residue F86^2.64, which appears to be important also for HEAT binding.     

Increased contribution of wheat nocturnal transpiration to daily water use under drought

Increasing evidence suggests that in crops, nocturnal water use could represent 30% of daytime water consumption, particularly in semi‐arid and arid areas. This raises the questions of whether nocturnal transpiration rates (TR_N) are (1) less influenced by drought than daytime TR (TR_D), (2) increased by higher nocturnal vapor pressure deficit (VPD_N), which prevails in such environments and (3) involved in crop drought tolerance. In this investigation, we addressed those questions by subjecting two wheat genotypes differing in drought tolerance to progressive soil drying under two long‐term VPD_N regimes imposed under naturally fluctuating conditions. A first goal was to characterize the response curves of whole‐plant TR_N and TR_N/TR_D ratios to progressive soil drying. A second goal was to examine the effect of VPD_N increase on TR_N response to soil drying and on 13 other developmental traits. The study revealed that under drought, TR_N was not responsive to progressive soil drying and – intriguingly – that TR_N seemingly increased with drought under high VPD_N consistently for the drought‐sensitive genotype. Because TR_D was concomitantly decreasing with progressive drought, this resulted in TR_N representing up to 70% of TR_D at the end of the drydown. In addition, under drought, VPD_N increase was found not to influence traits such as leaf area or stomata density. Overall, those findings indicate that TR_N contribution to daily water use under drought might be much higher than previously thought, that it is controlled by specific mechanisms and that decreasing TR_N under drought might be a valuable trait for improving drought tolerance.     

Do litter-mediated plant-soil feedbacks influence Mediterranean oak regeneration? A two-year pot experiment

Background & Aims

Oak seedling establishment is difficult and may be partly explained by litter-mediated interactions with neighbors. Litter effects can be physical or chemical and result in positive or negative feedback effects for seedlings. Mediterranean species leaves contain high levels of secondary metabolites which suggest that negative litter effects could be important.

Methods

Seedlings of Quercus ilex and Quercus pubescens were grown for two years in pots with natural soil and litter inputs from 6 Mediterranean woody species, artificial litter (only physical effect) or bare soil.

Results

Litter types had highly different mass loss (41–80%), which correlated with soil organic C, total N and microbial activity. Litter of Q. pubescens increased soil humidity and oak seedlings aerial biomass. Litters of Cotinus coggygria and Rosmarinus officinalis, containing high quantities of phenolics and terpenes respectively, decomposed fast and led to specific soil microbial catabolic profiles but did not influence oak seedling growth, chemistry or mycorrhization rates.

Conclusions

Physical litter effects through improved soil humidity seem to be predominant for oak seedling development. Despite high litter phenolics content, we detected no chemical effects on oak seedlings. Litter traits conferring a higher ability to retain soil moisture in dry periods deserve further attention as they may be critical to explain plant-soil feedbacks in Mediterranean ecosystems.

     

Reconstituting Corticostriatal Network on-a-Chip Reveals the Contribution of the Presynaptic Compartment to Huntington’s Disease

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