Hot-wiring dynein-2 establishes roles for IFT-A in retrograde train assembly and motility.

Intraflagellar transport (IFT) trains, built around IFT-A and IFT-B complexes, are carried by opposing motors to import and export ciliary cargo. While transported by kinesin-2 on anterograde IFT trains, the dynein-2 motor adopts an autoinhibitory conformation until it needs to be activated at the ciliary tip to power retrograde IFT. Growing evidence has linked the IFT-A complex to retrograde IFT; however, its roles in this process remain unknown. Here, we use CRISPR-Cas9-mediated genome editing to disable the dynein-2 autoinhibition mechanism in Caenorhabditis elegans and assess its impact on IFT with high-resolution live imaging and photobleaching analyses. Remarkably, this dynein-2 "hot-wiring" approach reignites retrograde motility inside IFT-A-deficient cilia without triggering tug-of-war events. In addition to providing functional evidence that multiple mechanisms maintain dynein-2 inhibited during anterograde IFT, our data establish key roles for IFT-A in mediating motor-train coupling during IFT turnaround, promoting retrograde IFT initiation, and modulating dynein-2 retrograde motility.

WDR60-mediated dynein-2 loading into cilia powers retrograde IFT and transition zone crossing.

The dynein-2 motor complex drives retrograde intraflagellar transport (IFT), playing a pivotal role in the assembly and functions of cilia. However, the mechanisms that regulate dynein-2 motility remain poorly understood. Here, we identify the Caenorhabditis elegans WDR60 homologue, WDR-60, and dissect the roles of this intermediate chain using genome editing and live imaging of endogenous dynein-2/IFT components. We find that loss of WDR-60 impairs dynein-2 recruitment to cilia and its incorporation onto anterograde IFT trains, reducing retrograde motor availability at the ciliary tip. Consistent with this, we show that fewer dynein-2 motors power WDR-60-deficient retrograde IFT trains, which move at reduced velocities and fail to exit cilia, accumulating on the distal side of the transition zone. Remarkably, disrupting the transition zone's NPHP module almost fully restores ciliary exit of underpowered retrograde trains in wdr-60 mutants. This work establishes WDR-60 as a major contributor to IFT, and the NPHP module as a roadblock to dynein-2 passage through the transition zone.

The IFT20 homolog in Caenorhabditis elegans is required for ciliogenesis and cilia-mediated behavior.

Cilia are microtubule-based organelles that carry out a wide range of critical functions throughout the development of higher animals. Regardless of their type, all cilia rely on a motor-driven, bidirectional transport system known as intraflagellar transport (IFT). Of the many components of the IFT machinery, IFT20 is one of the smallest subunits. Nevertheless, IFT20 has been shown to play critical roles in the assembly of several types of mammalian cilia. Here we show that the IFT20 homolog in Caenorhabditis elegans, IFT-20, is also important for correct cilium assembly in sensory neurons. Strikingly, however, we find that IFT-20-deficient animals are able to assemble short, vestigial cilia. In spite of this, we show that practically all IFT-20-deficient animals fail to respond to environmental cues that are normally detected by cilia to modulate their behavior. Altogether, our results indicate that IFT-20 is critical for both the correct assembly and function of cilia in C. elegans.

Characterization of New Proteomic Biomarker Candidates in Mucopolysaccharidosis Type IVA.

Mucopolysaccharidosis type IVA (MPS IVA) is a lysosomal storage disease caused by mutations in the N-acetylgalactosamine-6-sulfatase (GALNS) gene. Skeletal dysplasia and the related clinical features of MPS IVA are caused by disruption of the cartilage and its extracellular matrix, leading to a growth imbalance. Enzyme replacement therapy (ERT) with recombinant human GALNS has yielded positive results in activity of daily living and endurance tests. However, no data have demonstrated improvements in bone lesions and bone grow thin MPS IVA after ERT, and there is no correlation between therapeutic efficacy and urine levels of keratan sulfate, which accumulates in MPS IVA patients. Using qualitative and quantitative proteomics approaches, we analyzed leukocyte samples from healthy controls (n = 6) and from untreated (n = 5) and ERT-treated (n = 8, sampled before and after treatment) MPS IVA patients to identify potential biomarkers of disease. Out of 690 proteins identified in leukocytes, we selected a group of proteins that were dysregulated in MPS IVA patients with ERT. From these, we identified four potential protein biomarkers, all of which may influence bone and cartilage metabolism: lactotransferrin, coronin 1A, neutral alpha-glucosidase AB, and vitronectin. Further studies of cartilage and bone alterations in MPS IVA will be required to verify the validity of these proteins as potential biomarkers of MPS IVA.

Proteomic Analysis in Morquio A Cells Treated with Immobilized Enzymatic Replacement Therapy on Nanostructured Lipid Systems.

Morquio A syndrome, or mucopolysaccharidosis type IVA (MPS IVA), is a lysosomal storage disease due to mutations in the N-acetylgalactosamine-6-sulfatase (GALNS) gene. Systemic skeletal dysplasia and the related clinical features of MPS IVA are due to disruption of cartilage and its extracellular matrix, leading to an imbalance of growth. Enzyme replacement therapy (ERT) with recombinant human GALNS, alpha elosulfase, provides a systemic treatment. However, this therapy has a limited impact on skeletal dysplasia because the infused enzyme cannot penetrate cartilage and bone. Therefore, an alternative therapeutic approach to reach the cartilage is an unmet challenge. We have developed a new drug delivery system based on a nanostructure lipid carrier with the capacity to immobilize enzymes used for ERT and to target the lysosomes. This study aimed to assess the effect of the encapsulated enzyme in this new delivery system, using in vitro proteomic technology. We found a greater internalization of the enzyme carried by nanoparticles inside the cells and an improvement of cellular protein routes previously impaired by the disease, compared with conventional ERT. This is the first qualitative and quantitative proteomic assay that demonstrates the advantages of a new delivery system to improve the MPS IVA ERT.

Influence of resting energy expenditure on weight gain in adolescents taking second-generation antipsychotics.

BACKGROUND & AIMS: Weight gain is an undesirable side effect of second-generation antipsychotics (SGAs). We performed this study to examine the influence of SGAs on resting energy expenditure (REE) and the relationship of REE to weight gain in adolescent patients. METHODS: Antipsychotic-naïve or quasi-naïve (<72 h of exposure to antipsychotics) adolescent patients taking olanzapine, quetiapine, or risperidone in monotherapy were followed up for one year. We performed a prospective study (baseline, 1, 3, 6, and 12 months after treatment) based on anthropometric measurements, bioelectrical impedance analysis, and indirect calorimetry (Deltatrac™ II MBM-200) to measure REE. We also analyzed metabolic and hormonal data and adiponectin concentrations. RESULTS: Forty-six out of the 54 patients that started treatment attended at least 2 visits, and 16 completed 1 year of follow-up. Patients gained 10.8 ± 6.2 kg (60% in the form of fat mass) and increased their waist circumference by 11.1 ± 5.0 cm after 1 year of treatment. The REE/kg body mass ratio decreased (p = 0.027), and the REE/percentage fat-free mass (FFM) ratio increased (p = 0.007) following the fall in the percentage of FFM during treatment. Weight increase was significantly correlated with the REE/percentage FFM ratio at all the visits (1-3-6-12 months) (r = 0.69, p = 0.004 at 12 months). CONCLUSIONS: SGAs seem to induce a hypometabolic state (reflected as decreased REE/kg body mass and increased REE/percentage FFM). This could explain, at least in part, the changes in weight and body composition observed in these patients.

Attitude toward antipsychotic medication as a predictor of antipsychotic treatment discontinuation in first-episode early-onset psychosis.

BACKGROUND: Antipsychotic drug discontinuation is a key risk factor in psychotic relapses. Clinical relapse is related to poor outcome, especially in the earlier stages of psychotic illness. The attitude toward treatment during the acute phase of a first episode of psychosis has been proposed as one of the main determinants of treatment discontinuation. However, the relationship between attitude toward antipsychotic medication and treatment discontinuation in the adolescent population has not been properly assessed. METHODS: Adolescents, aged 12-18 years old, consecutively admitted to an adolescent unit with a first lifetime admission for a first episode of psychosis were asked to participate in a randomized, flexible-dose, 6-month controlled trial of olanzapine vs. quetiapine. Attitude toward antipsychotic medication was assessed using the 10-item Drug Attitude Inventory (DAI). The outcome variable was all-cause treatment discontinuation over the 6-month follow- up. The study sample was composed of 42 patients [34 boys (82.9%), eight girls (17.1%), mean age ± SD: 16.1±1.3]. RESULTS: Of the 42 patients, only 29 (69%) continued the medication throughout the entire 6-month follow-up, while 13 (31%) discontinued the medication. DAI scores were greater than zero at all assessments, indicating that the general attitude of the patients toward medication was positive. Higher DAI scores at baseline were related to lower all-cause treatment discontinuation [adjusted hazard ratio (HR) = 0.81 (95% CI: 0.68-0.96), P=0.016], while DAI scores at 15 days were unrelated to treatment discontinuation [adjusted HR=1.0 (95% CI: 0.82-1.23), P=0.998]. CONCLUSIONS: A better attitude toward antipsychotic medication at a first lifetime psychiatric admission for a first early-onset psychotic episode was significantly related to lower all-cause antipsychotic treatment discontinuation.

Does diagnostic classification of early-onset psychosis change over follow-up?

OBJECTIVE: To examine the diagnostic stability and the functional outcome of patients with early-onset psychosis (EOP) over a 2-year follow-up period. METHODS: A total of 24 patients (18 males (75%) and 6 females (25%), mean age +/- SD: 15.7 +/- 1.6 years) with a first episode of EOP formed the sample. Psychotic symptoms were assessed using the Positive and Negative Syndrome Scale (PANSS). Social disability was measured with the Global Assessment of Functioning (GAF) disability scale. Diagnosis was assessed using the Kiddie-Sads-Present and Lifetime Version (K-SADS-PL), according to DSM-IV criteria. All diagnoses were re-assessed after 1 year and 2 years. RESULTS: Schizophrenia had the highest prospective consistency (100% predictive value), while diagnostic stability was moderate for bipolar disorder (71.4%), and low for schizoaffective disorder (50%), schizophreniform and brief psychosis (50%), and psychosis not otherwise specified (NOS) (16.7%). The agreement between the baseline diagnoses and the 1-year follow-up diagnoses was 54.2%, whereas between the 1-year follow-up and the 2-year follow-up, it rose to 95.7%. Regardless of diagnosis, baseline negative symptoms were the only significant predictor of level of functioning at the 2-year follow-up (p = 0.010). CONCLUSION: Our results highlight the primacy of the first-year follow-up diagnosis in predicting diagnostic stability of EOP.