2-Bromopalmitate Reduces Protein Deacylation by Inhibition of Acyl-Protein Thioesterase Enzymatic Activities

S-acylation, the covalent attachment of palmitate and other fatty acids on cysteine residues, is a reversible post-translational modification that exerts diverse effects on protein functions. S-acylation is catalyzed by protein acyltransferases (PAT), while deacylation requires acyl-protein thioesterases (APT), with numerous inhibitors for these enzymes having already been developed and characterized. Among these inhibitors, the palmitate analog 2-brompalmitate (2-BP) is the most commonly used to inhibit palmitoylation in cells. Nevertheless, previous results from our laboratory have suggested that 2-BP could affect protein deacylation. Here, we further investigated in vivo and in vitro the effect of 2-BP on the acylation/deacylation protein machinery, with it being observed that 2-BP, in addition to inhibiting PAT activity in vivo, also perturbed the acylation cycle of GAP-43 at the level of depalmitoylation and consequently affected its kinetics of membrane association. Furthermore, 2-BP was able to inhibit in vitro the enzymatic activities of human APT1 and APT2, the only two thioesterases shown to mediate protein deacylation, through an uncompetitive mechanism of action. In fact, APT1 and APT2 hydrolyzed both the monomeric form as well as the micellar state of the substrate palmitoyl-CoA. On the basis of the obtained results, as APTs can mediate deacylation on membrane bound and unbound substrates, this suggests that the access of APTs to the membrane interface is not a necessary requisite for deacylation. Moreover, as the enzymatic activity of APTs was inhibited by 2-BP treatment, then the kinetics analysis of protein acylation using 2-BP should be carefully interpreted, as this drug also inhibits protein deacylation.     

Method for Quantitative Study of Airway Functional Microanatomy Using Micro-Optical Coherence Tomography

We demonstrate the use of a high resolution form of optical coherence tomography, termed micro-OCT (μOCT), for investigating the functional microanatomy of airway epithelia. μOCT captures several key parameters governing the function of the airway surface (airway surface liquid depth, periciliary liquid depth, ciliary function including beat frequency, and mucociliary transport rate) from the same series of images and without exogenous particles or labels, enabling non-invasive study of dynamic phenomena. Additionally, the high resolution of μOCT reveals distinguishable phases of the ciliary stroke pattern and glandular extrusion. Images and functional measurements from primary human bronchial epithelial cell cultures and excised tissue are presented and compared with measurements using existing gold standard methods. Active secretion from mucus glands in tissue, a key parameter of epithelial function, was also observed and quantified.     

Electrophysiological Heterogeneity of Fast-Spiking Interneurons: Chandelier versus Basket Cells

In the prefrontal cortex, parvalbumin-positive inhibitory neurons play a prominent role in the neural circuitry that subserves working memory, and alterations in these neurons contribute to the pathophysiology of schizophrenia. Two morphologically distinct classes of parvalbumin neurons that target the perisomatic region of pyramidal neurons, chandelier cells (ChCs) and basket cells (BCs), are generally thought to have the same “fast-spiking” phenotype, which is characterized by a short action potential and high frequency firing without adaptation. However, findings from studies in different species suggest that certain electrophysiological membrane properties might differ between these two cell classes. In this study, we assessed the physiological heterogeneity of fast-spiking interneurons as a function of two factors: species (macaque monkey vs. rat) and morphology (chandelier vs. basket). We showed previously that electrophysiological membrane properties of BCs differ between these two species. Here, for the first time, we report differences in ChCs membrane properties between monkey and rat. We also found that a number of membrane properties differentiate ChCs from BCs. Some of these differences were species-independent (e.g., fast and medium afterhyperpolarization, firing frequency, and depolarizing sag), whereas the differences in the first spike latency between ChCs and BCs were species-specific. Our findings indicate that different combinations of electrophysiological membrane properties distinguish ChCs from BCs in rodents and primates. Such electrophysiological differences between ChCs and BCs likely contribute to their distinctive roles in cortical circuitry in each species.     

Length-weight relationships of seven fish species from the laguna del cisne basin (Canelones,Uruguay)

This work provides new length-weight information for seven species of freshwater fishes. The analyzed specimens were collected monthly during a year (April 2018–March 2019) in a coastal lagoon system and its associated streams in southern Uruguay. During each sampling campaign, fishes were captured using gillnets in the lagoon and electrofishing in the streams. This study reports a new maximum size for four species and the first length-weight relationship report for Gymnogeophagus terrapurpura. Length-weight estimates and their confidence intervals are provided for all species.     

Polysome-associated lncRNAs during cardiomyogenesis of hESCs

Molecular and Cellular Biochemistry - In stress conditions, as neoplastic transformation, amino acids serve not only as nutrients to maintain the cell survival but also as mediators of several...     

Thermal ecology of two sympatric saxicolous lizards of the genus Phymaturus from the Payunia region (Argentina)

We evaluated the thermal biology of two sympatric saxicolous species of the genus Phymaturus, endemic from the Argentine Payunia region. Taking into account that the distributional range of Phymaturus roigorum (the largest species) is greater than the range of P. payuniae, we evaluated the habitat (type of rocks) used by these species. We recorded body temperature and operative temperatures in different habitats, and we determined the preferred body temperature in the laboratory. We compared the thermal quality of habitats occupied and not occupied by Phymaturus payuniae, and the accuracy and effectiveness of thermoregulation between species.     

Diagnostic utility of HFE variants in Spanish patients: Association with HLA alleles and role in susceptibility to acute lymphoblastic leukemia

Two single nucleotide polymorphisms (SNPs) in the Human Hemochromatosis (HFE) gene, C282Y and H63D, are the major variants associated to altered iron status and it is well known that these mutations are in linkage disequilibrium with certain Human Leukocyte Antigen (HLA)-A alleles. In addition, the C282Y SNP has been previously suggested to confer susceptibility to acute lymphoblastic leukemia (ALL). We have aimed to assess the diagnosis utility of these polymorphisms in a population of Spanish subjects with suspicion of hereditary iron overload and to evaluate the effect of their associations with HLA-A alleles on the susceptibility to ALL. Both the 63DD [OR = 4.31 (1.7–11.2)] and 282YY (p for trend = 0.02) genotypes were more frequently found among subjects with suspicion of iron overload than among controls. 282YY carriers displayed significantly higher transferrin saturation index (TSI) values (p < 0.001) as well as serum iron (p = 0.01) and ferritin (p = 0.01) levels. In addition, transferrin levels were lower in these subjects (p = 0.01). Likewise, patients who were carriers of the compound heterozygous diplotype (282CY/63HD) showed significantly higher TSI and serum iron and ferritin concentrations. The H63D SNP did not significantly affect the analytical parameters measured. All 282YY carriers and 69.2% of compound heterozygotes showed an altered biochemical index. The frequencies of the HFE SNPs in ALL pediatric patients were lower than those found in controls, whereas the HLA-A*24 allele was significantly overrepresented in the patients group [OR = 3.76 (1.9–7.3)]. No HFE-HLA-A associations were found to modulate the ALL risk. These results suggest that it may be useful to test for both HFE H63D and C282Y polymorphisms in patients with iron overload, as opposed to just genotyping for the C282Y SNP, which is customary in some healthcare centers. These HFE variants and their associations with HLA-A alleles were not observed to be relevant for the susceptibility to ALL in our population.     

Influence of MDR1 C1236T polymorphism on lopinavir plasma concentration and virological response in HIV-1-infected children

Background

Variability in MDR1 and PXR has been associated with differences in drug plasma levels and response to antiretroviral therapy. We investigated whether polymorphisms in MDR1 (T-129C, C1236T and C3435T) and PXR (C63396T) affect lopinavir plasma concentration and the virological or immunological response to HAART in HIV-1-infected children.

Methods

Genotypes were identified in 100 blood donors and 38 HIV-1-infected children. All children received HAART with lopinavir boosted with ritonavir (LPV/r) at the time of LPV plasma level quantification, before (C_trough) and between 1 and 2 h after (C_post-dose) the administration of the next dose of drug. CD4⁺ T-cell counts and plasma viral load were analyzed before and after the initiation of LPV/r.

Results

MDR1 1236T, MDR1 3435T and PXR 63396T alleles showed a frequency of ~ 50% while the MDR1 -129C allele only reached 5%. Children heterozygotes 1236CT showed a significantly lower LPV C_post-dose than homozygotes 1236TT (median C_post-dose = 3.04 μg/ml and 6.50 μg/ml, respectively; p = 0.016). Children heterozygotes 1236CT also had a lower decrease of viral load after 36 weeks of LPV/r exposure compared with homozygotes 1236CC (median viral load changes = − 0.50 log₁₀ copies/ml and − 2.08 log₁₀ copies/ml, respectively; p = 0.047). No effect on the immunological response was observed for polymorphisms of MDR1 or PXR.

Conclusions

Our results suggest that the MDR1 C1236T SNP significantly reduces LPV plasma concentration affecting the virological response to HAART. Heterozygotes 1236CT might have an altered level of P-gp expression/activity in enterocytes and CD4⁺ T lymphocytes that limits the absorption of LPV leading to an impaired virological suppression.