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Accumulating evidence indicates that G protein–coupled receptors (GPCRs) interact with Rab GTPases during their intracellular trafficking. How GPCRs recruit and activate the Rabs is unclear. Here, we report that depletion of endogenous L-type prostaglandin D synthase (L-PGDS) in HeLa cells inhibited recycling of the prostaglandin D(2) (PGD(2)) DP1 receptor (DP1) to the cell surface after agonist-induced internalization and that L-PGDS overexpression had the opposite effect. Depletion of endogenous Rab4 prevented l-PGDS–mediated recycling of DP1, and l-PGDS depletion inhibited Rab4-dependent recycling of DP1, indicating that both proteins are mutually involved in this pathway. DP1 stimulation promoted its interaction through its intracellular C terminus with Rab4, which was increased by l-PGDS. Confocal microscopy revealed that DP1 activation induces l-PGDS/Rab4 co-localization. l-PGDS/Rab4 and DP1/Rab4 co-immunoprecipitation levels were increased by DP1 agonist treatment. Pulldown assays with purified GST-l-PGDS and His(6)-Rab4 indicated that both proteins interact directly. l-PGDS interacted preferentially with the inactive, GDP-locked Rab4S22N variant rather than with WT Rab4 or with constitutively active Rab4Q67L proteins. Overexpression and depletion experiments disclosed that l-PGDS partakes in Rab4 activation following DP1 stimulation. Experiments with deletion mutants and synthetic peptides revealed that amino acids 85–92 in l-PGDS are involved in its interaction with Rab4 and in its effect on DP1 recycling. Of note, GTPγS loading and time-resolved FRET assays with purified proteins suggested that l-PGDS enhances GDP-GTP exchange on Rab4. Our results reveal how l-PGDS, which produces the agonist for DP1, regulates DP1 recycling by participating in Rab4 recruitment and activation.

Using a combination of neuroimaging and behavioural studies, Abdelgabar et al. show that the cerebellum helps us perceive the actions of others. Disorders such as spinocerebellar ataxia type 6, which disrupt cerebellar functioning, impair our ability to perceive the kinematics of other people’s actions, with potential implications for social cognition., Our cerebellum has been proposed to generate prediction signals that may help us plan and execute our motor programmes. However, to what extent our cerebellum is also actively involved in perceiving the action of others remains to be elucidated. Using functional MRI, we show here that observing goal-directed hand actions of others bilaterally recruits lobules VI, VIIb and VIIIa in the cerebellar hemispheres. Moreover, whereas healthy subjects (n = 31) were found to be able to discriminate subtle differences in the kinematics of observed limb movements of others, patients suffering from spinocerebellar ataxia type 6 (SCA6; n = 21) were severely impaired in performing such tasks. Our data suggest that the human cerebellum is actively involved in perceiving the kinematics of the hand actions of others and that SCA6 patients’ deficits include a difficulty in perceiving the actions of other individuals. This finding alerts us to the fact that cerebellar disorders can alter social cognition.

Aim: To comprehensively characterize the high-density lipoproteins (HDLs) microtranscriptome and to assess whether it is distinct from that of plasma and different between women and men. Methods: RNA was extracted from ultracentrifugation-purified HDLs and plasma from 17 healthy women and men couples, and libraries were sequenced on a HiSeq2500 platform. Results: On average, 310 ± 64 and 355 ± 31 miRNAs were detected (≥1 read per million) in HDLs and plasma, respectively. A total of 62 and 134 miRNAs were over-represented (e.g., miR-150-5p; fold change = 7.52; padj = 5.41 × 10-111) and under-represented (e.g., miR-22-3p; fold change = -5.28; padj = 2.11 × 10-154) in HDLs compared with plasma. These miRNAs were enriched in lipid metabolism and cellular processes-related pathways. Conclusion: HDLs exhibit a sex-independent miRNA profile distinct from that of plasma. These miRNAs may contribute to the HDLs' physiology.

Rab4a is a small GTPase associated with endocytic compartments and a key regulator of early endosomes recycling. Gathering evidence indicates that its expression and activation are required for the development of metastases. Rab4a-intrinsic GTPase properties that control its activity, i.e. nucleotide exchange and hydrolysis rates, have not yet been thoroughly studied. The determination of these properties is of the utmost importance to understand its functions and contributions to tumorigenesis. Here, we used the constitutively active (Rab4aQ67L) and dominant negative (Rab4aS22N) mutants to characterize the thermodynamical and structural determinants of the interaction between Rab4a and GTP (GTPγS) as well as GDP. We report the first 1H, 13C, 15N backbone NMR assignments of a Rab GTPase family member with Rab4a in complex with GDP and GTPγS. We also provide a qualitative description of the extent of structural and dynamical changes caused by the Q67L and S22N mutations. Using a real-time NMR approach and the two aforementioned mutants as controls, we evaluated Rab4a intrinsic nucleotide exchange and hydrolysis rates. Compared to most small GTPases such as Ras, a rapid GTP exchange rate along with slow hydrolysis rate were observed. This suggests that, in a cellular context, Rab4a can self-activate and persist in an activated state in absence of regulatory mechanisms. This peculiar profile is uncommon among the Ras superfamily members, making Rab4a an atypical fast-cycling GTPase and may explain, at least in part, how it contributes to metastases.

Our cerebellum has been proposed to generate prediction signals that may help us plan and execute our motor programs. However, to what extent our cerebellum is also actively involved in perceiving the action of others remains to be elucidated. Using fMRI, we show here that observing goal-directed hand actions of others bilaterally recruits cerebellar Lobules VI, VIIb and VIIIa. Moreover, whereas healthy subjects (n=31) were found to be able to discriminate subtle differences in the kinematics of observed limb movements of others, patients suffering from spinocerebellar ataxia type 6 (SCA6; n=21) were severely impaired in performing such tasks. Our data suggest that the human cerebellum is actively involved in perceiving the kinematics of the hand actions of others and that SCA6 patients’ deficits include a difficulty in perceiving the actions of other individuals. This finding alerts us to the fact that cerebellar disorders can alter social cognition. Given that impairments in social cognition have been reported to be one of the most debilitating consequences of neurological disorders, this finding may be relevant to improving the quality of life of patients and their families.

Many daily life activities demand precise integration of spatial and temporal information of sensory inputs followed by appropriate motor actions. This type of integration is carried out in part by the cerebellum, which has been postulated to play a central role in learning and timing of movements. Cerebellar damage due to atrophy or lesions may compromise forward-model processing, in which both spatial and temporal cues are used to achieve prediction for future motor states. In the present study we sought to further investigate the cerebellar contribution to predictive and reactive motor timing, as well as to learning of sequential order and temporal intervals in these tasks. We tested patients with spinocerebellar ataxia type 6 (SCA6) and healthy controls for two related motor tasks; one requiring spatio-temporal prediction of dynamic visual stimuli and another one requiring reactive timing only. We found that healthy controls established spatio-temporal prediction in their responses with high temporal precision, which was absent in the cerebellar patients. SCA6 patients showed lower predictive motor timing, coinciding with a reduced number of correct responses during the ‘anticipatory’ period on the task. Moreover, on the task utilizing reactive motor timing functions, control participants showed both sequence order and temporal interval learning, whereas patients only showed sequence order learning. These results suggest that SCA6 affects predictive motor timing and temporal interval learning. Our results support and highlight cerebellar contribution to timing and argue for cerebellar engagement during spatio-temporal prediction of upcoming events.