p24-Tango1 interactions ensure ER-Golgi interface stability and efficient transport.

The eukaryotic p24 family, consisting of α-, ß-, γ- and δ-p24 subfamilies, has long been known to be involved in regulating secretion. Despite increasing interest in these proteins, fundamental questions remain about their role. Here, we systematically investigated Drosophila p24 proteins. We discovered that members of all four p24 subfamilies are required for general secretion and that their localizations between ER exit site (ERES) and Golgi are interdependent in an α→ßδ→γ sequence. We also found that localization of p24 proteins and ERES determinant Tango1 requires interaction through their respective GOLD and SH3 lumenal domains, with Tango1 loss sending p24 proteins to the plasma membrane and vice versa. Finally, we show that p24 loss expands the COPII zone at ERES and increases the number of ER-Golgi vesicles, supporting a restrictive role of p24 proteins on vesicle budding for efficient transport. Our results reveal Tango1-p24 interplay as central to the generation of a stable ER-Golgi interface.

FGF signaling promotes spreading of fat body precursors necessary for adult adipogenesis in Drosophila.

Knowledge of adipogenetic mechanisms is essential to understand and treat conditions affecting organismal metabolism and adipose tissue health. In Drosophila, mature adipose tissue (fat body) exists in larvae and adults. In contrast to the well-known development of the larval fat body from the embryonic mesoderm, adult adipogenesis has remained mysterious. Furthermore, conclusive proof of its physiological significance is lacking. Here, we show that the adult fat body originates from a pool of undifferentiated mesodermal precursors that migrate from the thorax into the abdomen during metamorphosis. Through in vivo imaging, we found that these precursors spread from the ventral midline and cover the inner surface of the abdomen in a process strikingly reminiscent of embryonic mesoderm migration, requiring fibroblast growth factor (FGF) signaling as well. FGF signaling guides migration dorsally and regulates adhesion to the substrate. After spreading is complete, precursor differentiation involves fat accumulation and cell fusion that produces mature binucleate and tetranucleate adipocytes. Finally, we show that flies where adult adipogenesis is impaired by knock down of FGF receptor Heartless or transcription factor Serpent display ectopic fat accumulation in oenocytes and decreased resistance to starvation. Our results reveal that adult adipogenesis occurs de novo during metamorphosis and demonstrate its crucial physiological role.

JNK signaling and integrins cooperate to maintain cell adhesion during epithelial fusion in Drosophila.

The fusion of epithelial sheets is an essential and conserved morphogenetic event that requires the maintenance of tissue continuity. This is secured by membrane-bound or diffusible signals that instruct the epithelial cells, in a coordinated fashion, to change shapes and adhesive properties and when, how and where to move. Here we show that during Dorsal Closure (DC) in Drosophila, the Jun kinase (JNK) signaling pathway modulates integrins expression and ensures tissue endurance. An excess of JNK activity, as an outcome of a failure in the negative feedback implemented by the dual-specificity phosphatase Puckered (Puc), promotes the loss of integrins [the ß-subunit Myospheroid (Mys)] and amnioserosa detachment. Likewise, integrins signal back to the pathway to regulate the duration and strength of JNK activity. Mys is necessary for the regulation of JNK activity levels and in its absence, puc expression is downregulated and JNK activity increases.

Intrinsic and damage-induced JAK/STAT signaling regulate developmental timing by the Drosophila prothoracic gland.

Development involves tightly paced, reproducible sequences of events, yet it must adjust to conditions external to it, such as resource availability and organismal damage. A major mediator of damage-induced immune responses in vertebrates and insects is JAK/STAT signaling. At the same time, JAK/STAT activation by the Drosophila Upd cytokines is pleiotropically involved in normal development of multiple organs. Whether inflammatory and developmental JAK/STAT roles intersect is unknown. Here, we show that JAK/STAT is active during development of the prothoracic gland (PG), which controls metamorphosis onset through ecdysone production. Reducing JAK/STAT signaling decreased PG size and advanced metamorphosis. Conversely, JAK/STAT hyperactivation by overexpression of pathway components or SUMOylation loss caused PG hypertrophy and metamorphosis delay. Tissue damage and tumors, known to secrete Upd cytokines, also activated JAK/STAT in the PG and delayed metamorphosis, at least in part by inducing expression of the JAK/STAT target Apontic. JAK/STAT damage signaling, therefore, regulates metamorphosis onset by co-opting its developmental role in the PG. Our findings in Drosophila provide insights on how systemic effects of damage and cancer can interfere with hormonally controlled development and developmental transitions.

ER exit sites in Drosophila display abundant ER-Golgi vesicles and pearled tubes but no megacarriers.

Secretory cargos are collected at endoplasmic reticulum (ER) exit sites (ERES) before transport to the Golgi apparatus. Decades of research have provided many details of the molecular events underlying ER-Golgi exchanges. Essential questions, however, remain about the organization of the ER-Golgi interface in cells and the type of membrane structures mediating traffic from ERES. To investigate these, we use transgenic tagging in Drosophila flies, 3D-structured illumination microscopy (SIM), and focused ion beam scanning electron microscopy (FIB-SEM) to characterize ERES-Golgi units in collagen-producing fat body, imaginal discs, and imaginal discs overexpressing ERES determinant Tango1. Facing ERES, we find a pre-cis-Golgi region, equivalent to the vertebrate ER-Golgi intermediate compartment (ERGIC), involved in both anterograde and retrograde transport. This pre-cis-Golgi is continuous with the rest of the Golgi, not a separate compartment or collection of large carriers, for which we find no evidence. We observe, however, many vesicles, as well as pearled tubules connecting ERES and Golgi.

Katanin p60-like 1 sculpts the cytoskeleton in mechanosensory cilia.

Mechanoreceptor cells develop a specialized cytoskeleton that plays structural and sensory roles at the site of mechanotransduction. However, little is known about how the cytoskeleton is organized and formed. Using electron tomography and live-cell imaging, we resolve the 3D structure and dynamics of the microtubule-based cytoskeleton in fly campaniform mechanosensory cilia. Investigating the formation of the cytoskeleton, we find that katanin p60-like 1 (kat-60L1), a neuronal type of microtubule-severing enzyme, serves two functions. First, it amplifies the mass of microtubules to form the dense microtubule arrays inside the sensory cilia. Second, it generates short microtubules that are required to build the nanoscopic cytoskeleton at the mechanotransduction site. Additional analyses further reveal the functional roles of Patronin and other potential factors in the local regulatory network. In all, our results characterize the specialized cytoskeleton in fly external mechanosensory cilia at near-molecular resolution and provide mechanistic insights into how it is formed.

Non-Alcoholic Fatty Liver Disease Is Associated with Impairment of Ejection Fraction Among Individuals with Obesity Undergoing Bariatric Surgery: Results of a Cross-Sectional Study.

BACKGROUND: The relationship between non-alcoholic fatty liver disease (NAFLD) and myocardial function seems to be more than just the effect of mutual metabolic risk factors. OBJECTIVE: To determine whether there is a significant association between NAFLD assessed by means of liver biopsy and left ventricular function expressed by the estimated ejection fraction among individuals with obesity. METHODS: This is a cross-sectional study which enrolled individuals who consecutively underwent bariatric surgery. NAFLD was assessed by means of liver biopsies which were systematically collected during the procedures. The estimated ejection fraction was obtained by means of transthoracic echocardiograms. The main outcome evaluated was a possible association between NAFLD features and ejection fraction. The results of liver biopsies and the respective degrees of severity of each NAFLD feature were also correlated with the ejection fraction and main anthropometric, biochemical, and clinical variables. RESULTS: Of 112 individuals, 86.6% were female and the mean age was 38.5 ± 9.3 years. It was observed that the average estimated ejection fraction (EEF) was significantly lower among individuals with liver fibrosis (67.6 ± 5.5% vs. 70.8 ± 4.9%, p = 0.008). After adjustment for confounding variables in a multivariate model, the degree of liver fibrosis was independently associated with the EEF (R = - 0.3, p = 0.02). CONCLUSION: Among individuals with morbid obesity, the findings of this study are suggestive that liver fibrosis confirmed by histopathological examination is associated with a slight impairment of left ventricular function. Further studies are needed to confirm this association.

Renal Function 1 Year After Bariatric Surgery: Influence of Roux-en-Y Gastric Bypass and Identification of Pre-Operative Predictors of Improvement.

BACKGROUND: While evidence of improved renal function following gastric bypass exists, pre-operative predictors of this improvement are not completely known. OBJECTIVES: To assess the glomerular filtration rate (GFR) 1 year after Roux-en-Y gastric bypass (RYGB) and to identify pre-operative predictors associated with the improvement of renal function. METHODS: A historical cohort study, which included 109 obese patients before and 12 months after RYGB, was classified into subgroups according to GFR (normofiltration, hypofiltration (GFR < 5th percentile), and hyperfiltration (GFR > 95th percentile)). The 5th and 95th percentiles were 90 and 120 mL/min/1.73 m2, respectively. The primary outcome was the variation of GFR (%GFR) estimated by the Chronic Kidney Disease - Epidemiology Collaboration (CKD-EPI) formula, calculated using serum creatinine, ethnicity, and gender. RESULTS: The mean age was 38.3 ± 10.3 years and 77% were female; 52.3% presented hypertension and 27.5% type 2 diabetes. One year after surgery, the mean BMI decreased from 36.7 ± 3.6 to 28.8 ± 3.3 kg/m2 (p < 0.001). Pre-surgically, 37.6% presented hypofiltration, 47.7% normofiltration, and 14.7% hyperfiltration. The overall GFR increased from 95.5 ± 19 to 104 ± 16.4 mL/min (10.9%) (p < 0.001). The overall post-surgical %GFR was negatively correlated with the pre-surgical GFR (R = - 0.687; p < 0.001). In the hypofiltration and normofiltration subgroups, the post-surgical %GFR was negatively correlated with age (R = - 0.328, p = 0.036; and R = - 0.355, p = 0.004, respectively) and pre-surgical GFR (R = - 0.436, p = 0.04; and R = - 0.528, p < 0.001, respectively). CONCLUSION: RYGB led to a significant improvement in renal function, mainly among patients with a worse pre-operative renal function. In the hypofiltration and normofiltration subgroups, a younger age was associated with better outcomes.

Collagen secretion screening in Drosophila supports a common secretory machinery and multiple Rab requirements.

Collagens are large secreted trimeric proteins making up most of the animal extracellular matrix. Secretion of collagen has been a focus of interest for cell biologists in recent years because collagen trimers are too large and rigid to fit into the COPII vesicles mediating transport from the endoplasmic reticulum (ER) to the Golgi. Collagen-specific mechanisms to create enlarged ER-to-Golgi transport carriers have been postulated, including cargo loading by conserved ER exit site (ERES) protein Tango1. Here, we report an RNAi screening for genes involved in collagen secretion in Drosophila. In this screening, we examined distribution of GFP-tagged Collagen IV in live animals and found 88 gene hits for which the knockdown produced intracellular accumulation of Collagen IV in the fat body, the main source of matrix proteins in the larva. Among these hits, only two affected collagen secretion specifically: PH4αEFB and Plod, encoding enzymes known to mediate posttranslational modification of collagen in the ER. Every other intracellular accumulation hit affected general secretion, consistent with the notion that secretion of collagen does not use a specific mode of vesicular transport, but the general secretory pathway. Included in our hits are many known players in the eukaryotic secretory machinery, like COPII and COPI components, SNAREs and Rab-GTPase regulators. Our further analysis of the involvement of Rab-GTPases in secretion shows that Rab1, Rab2 and RabX3, are all required at ERES, each of them differentially affecting ERES morphology. Abolishing activity of all three by Rep knockdown, in contrast, led to uncoupling of ERES and Golgi. We additionally present a characterization of a screening hit we named trabuco (tbc), encoding an ERES-localized TBC domain-containing Rab-GAP. Finally, we discuss the success of our screening in identifying secretory pathway genes in comparison to two previous secretion screenings in Drosophila S2 cells.

Nationwide Macroeconomic Variables and the Growth Rate of Bariatric Surgeries in Brazil.

BACKGROUND: The effect of nationwide economic issues on the necessary expansion in the number of bariatric procedures remains unclear. OBJECTIVE: This study aims to determine whether there are correlations between the growth rate in the number of bariatric surgeries and the major macroeconomic variables over time in Brazil. METHODS: It is a nationwide analysis regarding the number of bariatric surgeries in Brazil and the main national macroeconomic variables from 2003 through 2016: gross domestic product (GDP), inflation rate, and the unemployment rate, as well as the evolution in the number of registered bariatric surgeons. RESULTS: There were significant positive correlations of the growth rate of surgeries with the early variations of the GDP (R = 0.5558; p = 0.04863) and of the overall health expenditure per capita (R = 0.78322; p = 0.00259). The growth rate of the number of bariatric surgeries was not correlated with the unemployment and inflation rates, as well as with the growth rate of available bariatric surgeons. CONCLUSION: There were direct relationships between the growth rate of bariatric surgeries and the evolutions of the GDP and health care expenditure per capita. These variables appear to influence the nationwide offer of bariatric surgery.