An Insulin-Responsive Sensor in the SIRT1 Disordered Region Binds DBC1 and PACS-2 to Control Enzyme Activity.

Current models of SIRT1 enzymatic regulation primarily consider the effects of fluctuating levels of its co-substrate NAD+, which binds to the stably folded catalytic domain. By contrast, the roles of the sizeable disordered N- and C-terminal regions of SIRT1 are largely unexplored. Here we identify an insulin-responsive sensor in the SIRT1 N-terminal region (NTR), comprising an acidic cluster (AC) and a 3-helix bundle (3HB), controlling deacetylase activity. The allosteric assistor DBC1 removes a distal N-terminal shield from the 3-helix bundle, permitting PACS-2 to engage the acidic cluster and the transiently exposed helix 3 of the 3-helix bundle, disrupting its structure and inhibiting catalysis. The SIRT1 activator (STAC) SRT1720 binds and stabilizes the 3-helix bundle, protecting SIRT1 from inhibition by PACS-2. Identification of the SIRT1 insulin-responsive sensor and its engagement by the DBC1 and PACS-2 regulatory hub provides important insight into the roles of disordered regions in enzyme regulation and the mode by which STACs promote metabolic fitness.

A Random Subcarrier-Selection Method Based on Index Modulation for Secure Transmission.

Recently, a frequency diverse array (FDA) has been employed in an orthogonal frequency division multiplexing (OFDM) transmitter to achieve secure wireless communication without mathematical encryption. However, an insecure coupling effect arises if the frequency increments are linearly assigned to all antenna elements. To solve this problem, random subcarrier-selection methods are proposed; however, the challenge lies in the random selection of subcarriers. Inspired by the randomness of index modulation (IM), this paper proposes a low complexity random subcarrier-selection method based on index modulation (RSCS-IM). Specifically, this work conducted analysis on the spectral efficiency (SE) of our system and the computational complexity of RSCS-IM, which works out a closed-form expression of the BER performance of a desired position and validates the theoretical outcomes through simulation.

Loss of carbonic anhydrase XII function in individuals with elevated sweat chloride concentration and pulmonary airway disease.

Elevated sweat chloride levels, failure to thrive (FTT), and lung disease are characteristic features of cystic fibrosis (CF, OMIM #219700). Here we describe variants in CA12 encoding carbonic anhydrase XII in two pedigrees exhibiting CF-like phenotypes. Exome sequencing of a white American adult diagnosed with CF due to elevated sweat chloride, recurrent hyponatremia, infantile FTT and lung disease identified deleterious variants in each CA12 gene: c.908-1 G>A in a splice acceptor and a novel frameshift insertion c.859_860insACCT. In an unrelated consanguineous Omani family, two children with elevated sweat chloride, infantile FTT, and recurrent hyponatremia were homozygous for a novel missense variant (p.His121Gln). Deleterious CFTR variants were absent in both pedigrees. CA XII protein was localized apically in human bronchiolar epithelia and basolaterally in the reabsorptive duct of human sweat glands. Respiratory epithelial cell RNA from the adult proband revealed only aberrant CA12 transcripts and in vitro analysis showed greatly reduced CA XII protein. Studies of ion transport across respiratory epithelial cells in vivo and in culture revealed intact CFTR-mediated chloride transport in the adult proband. CA XII protein bearing either p.His121Gln or a previously identified p.Glu143Lys missense variant localized to the basolateral membranes of polarized Madin-Darby canine kidney (MDCK) cells, but enzyme activity was severely diminished when assayed at physiologic concentrations of extracellular chloride. Our findings indicate that loss of CA XII function should be considered in individuals without CFTR mutations who exhibit CF-like features in the sweat gland and lung.

Population structure of Nouelia insignis (Asteraceae), an endangered species in southwestern China, based on chloroplast DNA sequences: recent demographic shrinking.

Nouelia insignis, an endangered species, is distributed in the Jinsha and Nanpan drainage areas in southwestern China. In this study, we examined the genetic diversity and population structure based on the sequences of the cpDNA rpL 16 intron. Low levels of genetic variation were detected within all populations of the endemic species. A gene genealogy of 11 haplotypes recovered two major lineages I and II, with haplotypes H1 and H6 nested as interior nodes, respectively. Haplotype H1 was widespread in all populations, while haplotype H6 was restricted to populations southern of the Jinsha River. Low levels of genetic differentiation were detected, as most F (st) values between populations were zero. This result, however, contradicts previous studies based on allozymes and fingerprinting. Genetic analyses suggested that coancestry due to low evolutionary rates resulted in the lack of geographical subdivision. Molecular dating estimated that the two lineages split about 3.224 MYA (95% CI 1.070-6.089 MYA). Maintenance of ancestral polymorphisms was possibly attributable to a long-standing large effective population size until recently. Postglacial demographic expansion was supported by a unimodal mismatch distribution and star-like phylogenies.

The multifunctional protein PACS-1 is required for HDAC2- and HDAC3-dependent chromatin maturation and genomic stability.

Phosphofurin acidic cluster sorting protein-1 (PACS-1) is a multifunctional membrane traffic regulator that plays important roles in organ homeostasis and disease. In this study, we elucidate a novel nuclear function for PACS-1 in maintaining chromosomal integrity. PACS-1 progressively accumulates in the nucleus during cell cycle progression, where it interacts with class I histone deacetylases 2 and 3 (HDAC2 and HDAC3) to regulate chromatin dynamics by maintaining the acetylation status of histones. PACS-1 knockdown results in the proteasome-mediated degradation of HDAC2 and HDAC3, compromised chromatin maturation, as indicated by elevated levels of histones H3K9 and H4K16 acetylation, and, consequently, increased replication stress-induced DNA damage and genomic instability.

Triterpenoids from Ganoderma lucidum and Their Potential Anti-inflammatory Effects.

Ganoderma lucidum, as food, tea, dietary supplement, and medicine, is widely used in China and Eastern Asian countries. In order to discover its anti-inflammatory constituents and provide some references for the usage of G. lucidum and G. sinense, two official species in China, the fruiting bodies of G. lucidum were studied, leading to the isolation of six new triterpenoids (1-6) and 27 known analogues (7-33). Compound 4 exhibited the most potent inhibition on nitric oxide (NO) production induced by lipopolysaccharide (LPS) in RAW264.7 macrophage cells. The production of IL-6 and IL-1ß, as well as the expression of iNOS, COX-2, and NF-κB were dose-dependently reduced by 4. The phosphorylations of IκBα and IKKß in LPS-induced macrophage cells were blocked by 4. Therefore, 4 could be used as a potential anti-inflammatory candidate and the total triterpenoids might be developed as value-added functional food for the prevention of inflammation. In combination of previous studies, it should be cautious for the interchangeable usage of G. lucidum and G. sinense.

Labdanes and megastigmanes from Vitex negundo var. heterophylla.

The aromatic plants of Vitex negundo var. heterophylla are not only herb medicine but also a functional food and an industrial crop. Its leaves can be used as a functional food for improving human's health, but the previous studies mainly focused on the volatile constituents, lignans, and iridoids. Our research led to the isolation of four new terpenoids (1-4), together with fifteen known compounds including seven flavonoids (9-15), two jasmonates (7-8) and six terpenoids (5-6, 16-19) from the leaves. Among all these compounds, 1, 2, 11, and 19 exhibited strong inhibitory activity against NO production in lipopolysaccharide (LPS)-induced RAW264.7 macrophage. The anti-inflammatory mechanism of the most active compound (2) is related to the inhibition of iNOS and COX-2, and the suppression of NF-κB pathway. Therefore, terpenoids and flavonoids from the leaves of Vitex negundo var. heterophylla might be used as potential anti-inflammatory candidates for developing medicine or value-added functional food.

The lysosomal enzyme receptor protein (LERP) is not essential, but is implicated in lysosomal function in Drosophila melanogaster.

The lysosomal enzyme receptor protein (LERP) of Drosophila melanogaster is the ortholog of the mammalian cation-independent mannose 6-phosphate (Man 6-P) receptor, which mediates trafficking of newly synthesized lysosomal acid hydrolases to lysosomes. However, flies lack the enzymes necessary to make the Man 6-P mark, and the amino acids implicated in Man 6-P binding by the mammalian receptor are not conserved in LERP. Thus, the function of LERP in sorting of lysosomal enzymes to lysosomes in Drosophila is unclear. Here, we analyze the consequence of LERP depletion in S2 cells and intact flies. RNAi-mediated knockdown of LERP in S2 cells had little or no effect on the cellular content or secretion of several lysosomal hydrolases. We generated a novel Lerp null mutation, Lerp(F6), which abolishes LERP protein expression. Lerp mutants have normal viability and fertility and display no overt phenotypes other than reduced body weight. Lerp mutant flies exhibit a 30-40% decrease in the level of several lysosomal hydrolases, and are hypersensitive to dietary chloroquine and starvation, consistent with impaired lysosome function. Loss of LERP also enhances an eye phenotype associated with defective autophagy. Our findings implicate Lerp in lysosome function and autophagy.

The unique GGA clathrin adaptor of Drosophila melanogaster is not essential.

The Golgi-localized, γ-ear-containing, ARF binding proteins (GGAs) are a highly conserved family of monomeric clathrin adaptor proteins implicated in clathrin-mediated protein sorting between the trans-Golgi network and endosomes. GGA RNAi knockdowns in Drosophila have resulted in conflicting data concerning whether the Drosophila GGA (dGGA) is essential. The goal of this study was to define the null phenotype for the unique Drosophila GGA. We describe two independently derived dGGA mutations. Neither allele expresses detectable dGGA protein. Homozygous and hemizygous flies with each allele are viable and fertile. In contrast to a previous report using RNAi knockdown, GGA mutant flies show no evidence of age-dependent retinal degeneration or cathepsin missorting. Our results demonstrate that several of the previous RNAi knockdown phenotypes were the result of off-target effects. However, GGA null flies are hypersensitive to dietary chloroquine and to starvation, implicating GGA in lysosomal function and autophagy.

Determination of DNA methylation of imprinted genes in Arabidopsis endosperm.

Arabidopsis thaliana is an excellent model organism for studying epigenetic mechanisms. One of the reasons is the loss-of-function null mutant of DNA methyltransferases is viable, thus providing a system to study how loss of DNA methylation in a genome affects growth and development. Imprinting refers to differential expression of maternal and paternal alleles and plays an important role in reproduction development in both mammal and plants. DNA methylation is critical for determining whether the maternal or paternal alleles of an imprinted gene is expressed or silenced. In flowering plants, there is a double fertilization event in reproduction: one sperm cell fertilizes the egg cell to form embryo and a second sperm fuses with the central cell to give rise to endosperm. Endosperm is the tissue where imprinting occurs in plants. MEDEA, a SET domain Polycomb group gene, and FWA, a transcription factor regulating flowering, are the first two genes shown to be imprinted in endosperm and their expression is controlled by DNA methylation and demethylation in plants. In order to determine imprinting status of a gene and methylation pattern in endosperm, we need to be able to isolate endosperm first. Since seed is tiny in Arabidopsis, it remains challenging to isolate Arabidopsis endosperm and examine its methylation. In this video protocol, we report how to conduct a genetic cross, to isolate endosperm tissue from seeds, and to determine the methylation status by bisulfite sequencing.