Defining the Essential Function of Yeast Hsf1 Reveals a Compact Transcriptional Program for Maintaining Eukaryotic Proteostasis.

Despite its eponymous association with the heat shock response, yeast heat shock factor 1 (Hsf1) is essential even at low temperatures. Here we show that engineered nuclear export of Hsf1 results in cytotoxicity associated with massive protein aggregation. Genome-wide analysis revealed that Hsf1 nuclear export immediately decreased basal transcription and mRNA expression of 18 genes, which predominately encode chaperones. Strikingly, rescuing basal expression of Hsp70 and Hsp90 chaperones enabled robust cell growth in the complete absence of Hsf1. With the exception of chaperone gene induction, the vast majority of the heat shock response was Hsf1 independent. By comparative analysis of mammalian cell lines, we found that only heat shock-induced but not basal expression of chaperones is dependent on the mammalian Hsf1 homolog (HSF1). Our work reveals that yeast chaperone gene expression is an essential housekeeping mechanism and provides a roadmap for defining the function of HSF1 as a driver of oncogenesis.

SMARCE1 is required for the invasive progression of in situ cancers.

Advances in mammography have sparked an exponential increase in the detection of early-stage breast lesions, most commonly ductal carcinoma in situ (DCIS). More than 50% of DCIS lesions are benign and will remain indolent, never progressing to invasive cancers. However, the factors that promote DCIS invasion remain poorly understood. Here, we show that SMARCE1 is required for the invasive progression of DCIS and other early-stage tumors. We show that SMARCE1 drives invasion by regulating the expression of secreted proteases that degrade basement membrane, an ECM barrier surrounding all epithelial tissues. In functional studies, SMARCE1 promotes invasion of in situ cancers growing within primary human mammary tissues and is also required for metastasis in vivo. Mechanistically, SMARCE1 drives invasion by forming a SWI/SNF-independent complex with the transcription factor ILF3. In patients diagnosed with early-stage cancers, SMARCE1 expression is a strong predictor of eventual relapse and metastasis. Collectively, these findings establish SMARCE1 as a key driver of invasive progression in early-stage tumors.

Quantitative Neurotoxicology: An Assessment of the Neurotoxic Profile of Kainic Acid in Sprague Dawley Rats.

This study consisted of a qualitative and quantitative assessment of neuropathological changes in kainic acid (KA)-treated adult male rats. Rats were administered a single 10 mg/kg intraperitoneal injection of KA or the same volume of saline and sacrificed 24 or 48 hours posttreatment. Brains were collected, sectioned coronally (∼ 81 slices), and stained with amino cupric silver to reveal degenerative changes. For qualitative assessment of neural degeneration, sectioned material was evaluated by a board-certified pathologist, and the level of degeneration was graded based upon a 4-point scale. For measurement of quantitative neural degeneration in response to KA treatment, the HALO digital image analysis software tool was used. Quantitative measurements of specific regions within the brain were obtained from silver-stained tissue sections with quantitation based on stain color and optical density. This quantitative evaluation method identified degeneration primarily in the cerebral cortex, septal nuclei, amygdala, olfactory bulb, hippocampus, thalamus, and hypothalamus. The KA-produced neuronal degeneration in the cortex was primarily in the piriform, insular, rhinal, and cingulate areas. In the hippocampus, the dentate gyrus was found to be the most affected area. Our findings indicate global neurotoxicity due to KA treatment. Certain brain structures exhibited more degeneration than others, reflecting differential sensitivity or vulnerability of neurons to KA.

GSK-3ß Phosphorylation of Cytoplasmic Dynein Reduces Ndel1 Binding to Intermediate Chains and Alters Dynein Motility.

Glycogen synthase kinase 3 (GSK-3) has been linked to regulation of kinesin-dependent axonal transport in squid and flies, and to indirect regulation of cytoplasmic dynein. We have now found evidence for direct regulation of dynein by mammalian GSK-3ß in both neurons and non-neuronal cells. GSK-3ß coprecipitates with and phosphorylates mammalian dynein. Phosphorylation of dynein intermediate chain (IC) reduces its interaction with Ndel1, a protein that contributes to dynein force generation. Two conserved residues, S87/T88 in IC-1B and S88/T89 in IC-2C, have been identified as GSK-3 targets by both mass spectrometry and site-directed mutagenesis. These sites are within an Ndel1-binding domain, and mutation of both sites alters the interaction of IC's with Ndel1. Dynein motility is stimulated by (i) pharmacological and genetic inhibition of GSK-3ß, (ii) an insulin-sensitizing agent (rosiglitazone) and (iii) manipulating an insulin response pathway that leads to GSK-3ß inactivation. Thus, our study connects a well-characterized insulin-signaling pathway directly to dynein stimulation via GSK-3 inhibition.

A Cdk5-dependent switch regulates Lis1/Ndel1/dynein-driven organelle transport in adult axons.

Lissencephaly is a human developmental brain abnormality caused by LIS1 haploinsufficiency. This disorder is in large part attributed to altered mitosis and migration in the developing brain. LIS1 and an interacting protein, NDEL1, bind to cytoplasmic dynein, a microtubule motor protein. While the tripartite complex is clearly important for developmental events, we are intrigued by the fact that Lis1 and Ndel1 expression remain high in the adult mouse nervous system. Dynein plays a crucial role in retrograde axonal transport, a process that is used by mature neurons. Here, we monitored acidic organelles moving in axons of adult rat sensory neurons to determine whether Lis1 and Ndel1 contribute to axonal transport. Lis1 RNAi significantly reduced axon transport of these organelles. Ndel1 RNAi had little impact, but combined Lis1 and Ndel1 RNAi caused a more severe phenotype than Lis1 RNAi alone, essentially shutting down transport. Lis1 overexpression stimulated retrograde transport, while a Lis1 dynein-binding mutant severely disrupted transport. Overexpression of Ndel1 or a Lis1 Ndel1-binding mutant only mildly perturbed transport. However, expressing a mutant Ndel1 lacking key phosphorylation sites shut down transport completely, as did a dominant-negative Cdk5 construct. We propose that, in axons, unphosphorylated Ndel1 inhibits the capacity of dynein to transport acidic organelles. Phosphorylation of Ndel1 by Cdk5 not only reduces this inhibition but also allows Lis1 to further stimulate the cargo transport capacity of dynein. Our data raise the possibility that defects in a Lis1/Ndel1 regulatory switch could contribute to neurodegenerative diseases linked to axonal pathology in adults.

LIS1 and NDEL1 Regulate Axonal Trafficking of Mitochondria in Mature Neurons.

Defective mitochondrial dynamics in axons have been linked to both developmental and late-onset neurological disorders. Axonal trafficking is in large part governed by the microtubule motors kinesin-1 and cytoplasmic dynein 1 (dynein). Dynein is the primary retrograde transport motor in axons, and mutations in dynein and many of its regulators also cause neurological diseases. Depletion of LIS1, famous for linking dynein deregulation to lissencephaly (smooth brain), in adult mice leads to severe neurological phenotypes, demonstrating post-developmental roles. LIS1 stimulates retrograde transport of acidic organelles in cultured adult rat dorsal root ganglion (DRG) axons but findings on its role in mitochondrial trafficking have been inconsistent and have not been reported for adult axons. Here we report that there is an increased number of mitochondria in cross-sections of sciatic nerve axons from adult LIS1+/- mice. This is probably related to reduced dynein activity as axons from adult rat nerves exposed to the dynein inhibitor, ciliobrevin D also had increased numbers of mitochondria. Moreover, LIS1 overexpression (OE) in cultured adult rat DRG axons stimulated retrograde mitochondrial transport while LIS1 knockdown (KD) or expression of a LIS1 dynein-binding mutant (LIS1-K147A) inhibited retrograde transport, as did KD of dynein heavy chain (DHC). These findings are consistent with our report on acidic organelles. However, KD of NDEL1, a LIS1 and dynein binding protein, or expression of a LIS1 NDEL1-binding mutant (LIS1-R212A) also dramatically impacted retrograde mitochondrial transport, which was not the case for acidic organelles. Manipulations that disrupted retrograde mitochondrial transport also increased the average length of axonal mitochondria, suggesting a role for dynein in fusion or fission events. Our data point to cargo specificity in NDEL1 function and raise the possibility that defects in the LIS1/NDEL1 dynein regulatory pathway could contribute to mitochondrial diseases with axonal pathologies.

Carbon footprint: current methods of estimation.

Increasing greenhouse gaseous concentration in the atmosphere is perturbing the environment to cause grievous global warming and associated consequences. Following the rule that only measurable is manageable, mensuration of greenhouse gas intensiveness of different products, bodies, and processes is going on worldwide, expressed as their carbon footprints. The methodologies for carbon footprint calculations are still evolving and it is emerging as an important tool for greenhouse gas management. The concept of carbon footprinting has permeated and is being commercialized in all the areas of life and economy, but there is little coherence in definitions and calculations of carbon footprints among the studies. There are disagreements in the selection of gases, and the order of emissions to be covered in footprint calculations. Standards of greenhouse gas accounting are the common resources used in footprint calculations, although there is no mandatory provision of footprint verification. Carbon footprinting is intended to be a tool to guide the relevant emission cuts and verifications, its standardization at international level are therefore necessary. Present review describes the prevailing carbon footprinting methods and raises the related issues.

Modeling and Optimization of Process Parameters for Nutritional Enhancement in Enzymatic Milled Rice by Multiple Linear Regression (MLR) and Artificial Neural Network (ANN).

This study involves information about the concentrations of nutrients (proteins, phenolic compounds, free amino acids, minerals (Ca, P, and Iron), hardness) in milled rice processed with enzymes; xylanase and cellulase produced by Aspergillus awamori, MTCC 9166 and Trichoderma reese, MTCC164. Brown rice was processed with 60-100% enzyme (40 mL buffer -undiluted) for 30 to 150 min at 30 °C to 50 °C followed by polishing for 20-100 s at a safe moisture level. Multiple linear regression (MLR) and artificial neural network (ANN) models were used for process optimization of enzymes. The MLR correlation coefficient (R2) varied between 0.87-0.90, and the sum of square (SSE) was placed within 0.008-8.25. While the ANN R2 (correlation coefficient) varied between 0.97 and 0.9999(1), MSE changed from 0.005 to 6.13 representing that the ANN method has better execution across MLR. The optimized cellulase process parameters (87.2% concentration, 80.1 min process time, 33.95 °C temperature and 21.8 s milling time) and xylanase process parameters (85.7% enzyme crude, 77.1 min process time, 35 °C temperature and 20 s) facilitated the increase of Ca (70%), P (64%), Iron (17%), free amino acids (34%), phenolic compounds (78%) and protein (84%) and decreased hardness (20%) in milled rice. Scanning electron micrographs showed an increased rupture attributing to enzymes action on milled rice.

Critical overview of biomass feedstocks as sustainable substrates for the production of polyhydroxybutyrate (PHB).

Polyhydroxybutyrates (PHBs) are a class of biopolymers produced by different microbial species and are biodegradable and biocompatible in nature as opposed to petrochemically derived plastics. PHBs have advanced applications in medical sector, packaging industries, nanotechnology and agriculture, among others. PHB is produced using various feedstocks such as glycerol, dairy wastes, agro-industrial wastes, food industry waste and sugars. Current focus on PHB research has been primarily on reducing the cost of production and, on downstream processing to isolate PHB from cells. Recent advancements to improve the productivity and quality of PHB include genetic modification of producer strain and modification of PHB by blending to develop desirable properties suited to diversified applications. Selection of feedstock plays a critical role in determining the economic feasibility and sustainability of the process. This review provides a bird's eye view of the suitability of different waste resources for producing polyhydroxybutyrate; providing state-of the art information and analysis.