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Conditional knock-out

Bryan E McGill, Ruteja A Barve, Susan E Maloney, Amy Strickland, Nicholas Rensing, Peter Wang, Michael Wong, Richard Head, David F Wozniak, Jeffrey Milbrandt
CCCTC-binding factor (CTCF) is an 11 zinc finger DNA-binding domain protein that regulates gene expression by modifying three dimensional chromatin structure. Human mutations in CTCF cause intellectual disability and autistic features. Knocking out Ctcf in mouse embryonic neurons is lethal by neonatal age, but the effects of CTCF deficiency in postnatal neurons are less well studied. We knocked out Ctcf postnatally in glutamatergic forebrain neurons under the control of Camk2a-CreCtcf(loxP/loxP) ;Camk2a-Cre+ (Ctcf CKO) mice of both sexes were viable and exhibited profound deficits in spatial learning/memory, impaired motor co-ordination, and decreased sociability by 4 months of age...
November 13, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
Shabina Parveen, Atish Roy Chowdhury, Junaid Jibran Jawed, Suchandra Bhattacharyya Majumdar, Bhaskar Saha, Subrata Majumdar
DUSP4, an inducible protein has a substrate specificity toward ERK1/2, a component of MAP kinase which is enhanced during Leishmania infection. The DUSP4(-/-) mice show increased susceptibility towards the infection caused by Toxoplasma gondii and Leishmania mexicana. These observations emphatically established the fact that unlike DUSP1, DUSP4 has host protective role. In our study, it has been Leishmania donovani, the causative agent of visceral leishmaniasis (VL) significantly reduced the expression of DUSP4 during infection...
November 10, 2017: Microbes and Infection
Clara Ballesté-Delpierre, Dietmar Fernandez-Orth, Mario Ferrer-Navarro, Ramón Díaz-Peña, Antonia Odena-Caballol, Eliandre Oliveira, Anna Fàbrega, Jordi Vila
Salmonella possesses virulence determinants that allow replication under extreme conditions and invasion of host cells, causing disease. Here, we examined four putative genes predicted to encode membrane proteins (ydiY, ybdJ, STM1441 and ynaJ) and a putative transcriptional factor (yedF). These genes were identified in a previous study of a S. Typhimurium clinical isolate and its multidrug-resistant counterpart. For STM1441 and yedF a reduced ability to interact with HeLa cells was observed in the knock-out mutants, but an increase in this ability was absent when these genes were overexpressed, except for yedF which phenotype was rescued when yedF was restored...
November 10, 2017: Scientific Reports
Andreas J Genewsky, Carsten T Wotjak
Among the hardwired behaviors, fear or survival responses certainly belong to the most evolutionary conserved ones. However, higher animals possess the ability to adapt to certain environments (e.g., novel foraging grounds), and, therefore, those responses need to be plastic. Previous studies revealed a cell-type specific role of the endocannabinoid system in novelty fear, conditioned fear and active vs. passive avoidance in a shuttle box paradigm. In this study we aim to investigate, whether knocking-out the cannabinoid receptor type-1 (CB1) on cortical glutamatergic (Glu-CB1(-/-)) or GABAergic (GABA-CB1(-/-)) neurons differentially affects the level of behavioral inhibition, which could ultimately lead to differences in escape behavior...
2017: Frontiers in Behavioral Neuroscience
Aiden Eblimit, Smriti Akshay Agrawal, Kandace Thomas, Ivan Assenov Anastassov, Tajiguli Abulikemu, Graeme Mardon, Rui Chen
The mammalian retina consists of multiple cell layers including photoreceptor cells, which are light sensing neurons that play essential functions in the visual process. Previously, we identified mutations in SPATA7, encoding spermatogenesis associated protein 7, in families with Leber Congenital Amaurosis (LCA) and juvenile Retinitis Pigmentosa (RP), and showed that Spata7 null mice recapitulate the human disease phenotype of retinal degeneration. SPATA7 is expressed in the connecting cilium of photoreceptor (PR) cells in the mouse retina, as well as in retinal pigment epithelium (RPE) cells, but the functional role of Spata7 in the RPE remains unknown...
October 31, 2017: Experimental Eye Research
Li Tang, Bigang Mao, Yaokui Li, Qiming Lv, LiPing Zhang, Caiyan Chen, Hanjie He, Weiping Wang, Xiongfeng Zeng, Ye Shao, Yinlin Pan, Yuanyi Hu, Yan Peng, Xiqin Fu, Hongqing Li, Shitou Xia, Bingran Zhao
Rice grain with excessive cadmium (Cd) is a major source of dietary Cd intake and a serious threat to health for people who consume rice as a staple food. The development of elite rice cultivars with consistently low Cd content is challenging for conventional breeding approaches, and new strategies urgently need to be developed. Here, we report the development of new indica rice lines with low Cd accumulation and no transgenes by knocking out the metal transporter gene OsNramp5 using CRISPR/Cas9 system. Hydroponic culture showed that Cd concentrations in shoots and roots of osnramp5 mutants were dramatically decreased, resulting in rescue of impaired growth in high Cd condition...
October 31, 2017: Scientific Reports
Kristina Zaprazna, Arindam Basu, Nikola Tom, Vibha Jha, Suchita Hodawadekar, Lenka Radova, Jitka Malcikova, Boris Tichy, Sarka Pospisilova, Michael L Atchison
Activation-induced cytidine deminase (AID) is crucial for controlling the immunoglobulin (Ig) diversification processes of somatic hypermutation (SHM) and class switch recombination (CSR). AID initiates these processes by deamination of cytosine, ultimately resulting in mutations or double strand DNA breaks needed for SHM and CSR. Levels of AID control mutation rates, and off-target non-Ig gene mutations can contribute to lymphomagenesis. Therefore, factors that control AID levels in the nucleus can regulate SHM and CSR, and may contribute to disease...
October 28, 2017: European Journal of Immunology
Tanner Byer, Jessica Wang, Mark G Zhang, Naomi Vather, Anna Blachman, Bryan Visser, Jane M Liu
The phosphoenopyruvate:carbohydrate phosphotransferase system (PTS) enables Vibrio cholerae - and other bacteria - to recognize and transport exogenous carbon sources for energy, including the six-carbon sugar alcohol, mannitol. The mannitol-specific PTS transporter is encoded by mtlA and its expression is expected to be regulated by the putative repressor encoded by the mtlR gene. Here, we show that mtlR overexpression inhibits V. cholerae growth in medium supplied with mannitol as the sole carbon source and represses MtlA-mediated biofilm formation...
October 27, 2017: Microbiology
Weiwei Gao, Runxia Shao, Xiaoping Zhang, Daijian Liu, Ying Liu, Xian'en Fa
Pulmonary arterial hypertension (PAH), characterized by excessive proliferation and apoptosis resistance of pulmonary artery smooth muscle cells (PASMCs), is closely associated with the imbalance in vasoactive mediators and massive remodeling of pulmonary vasculature. DJ-1/park7, a multifunctional protein, plays a critical defense role in several cytobiological activity, such as transcriptional regulation, anti-oxidative stress and tumor formation. In this study, we investigated the effects of DJ-1 on hypoxia-induced PAH model rats and PASMCs, as well as its possible molecular mechanism...
October 22, 2017: Experimental Cell Research
Natalia Madrid-López, Jorge Estrada, Javier Díaz, Alejandro Bassi, Paul H Délano, Adrián Ocampo-Garcés
There is a neural matrix controlling the sleep-wake cycle (SWC) embedded within high ranking integrative mechanisms in the central nervous system. Nicotinic alpha-9 acetylcholine receptor subunit (alpha-9 nAChR) participate in physiological processes occurring in sensory, endocrine and immune systems. There is a relationship between the SWC architecture, body homeostasis and sensory afferents so that disruption of afferent signaling is expected to affect the temporal organization of sleep and wake states. The analysis of the SWC of 9 nAChR knock-out animals may help to reveal the contribution of alpha-9 nAChR to sleep chronobiological determinants...
2017: Frontiers in Cellular Neuroscience
Lempie Kashinasha Ekandjo, Silke Ruppel, Rainer Remus, Katja Witzel, Sascha Patz, Yvonne Becker
Biological nitrogen fixation (BNF) is considered one of the key plant growth-promoting (PGP) factors for diazotrophic organisms. Whether the Fe and the FeMo nitrogenases of <i>Kosakonia radicincitans</i> contribute to its PGP effect is yet to be proven. Hence, for the first time we conducted site-directed mutagenesis in <i>K. radicincitans</i> to knock out <i>anfH</i> and/or <i>nifH</i> as means to deactivate BNF in this strain. We used <sup>15</sup>N<sub>2</sub> labeled air to trace BNF activities in Δ<i>anfH</i> Δ<i>nifH</i> and Δ<i>anfH</i>Δ<i>nifH</i> mutants...
October 23, 2017: Canadian Journal of Microbiology
Michele Conroy, Jennifer A Soltis, Rick S Wittman, Frances N Smith, Sayandev Chatterjee, Xin Zhang, Eugene S Ilton, Edgar C Buck
Layered (oxy) hydroxide minerals often possess out-of-plane hydrogen atoms that form hydrogen bonding networks which stabilize the layered structure. However, less is known about how the ordering of these bonds affects the structural stability and solubility of these minerals. Here, we report a new strategy that uses the focused electron beam to probe the effect of differences in hydrogen bonding networks on mineral solubility. In this regard, the dissolution behavior of boehmite (γ-AlOOH) and gibbsite (γ-Al(OH)3) were compared and contrasted in real time via liquid cell electron microscopy...
October 16, 2017: Scientific Reports
Valérie Bergeron, Julien Ghislain, Kevin Vivot, Natalia Tamarina, Louis H Philipson, Jens Fielitz, Vincent Poitout
Beta-cell adaptation to insulin resistance is necessary to maintain glucose homeostasis in obesity. Failure of this mechanism is a hallmark of type 2 diabetes (T2D). Hence, factors controlling functional beta-cell compensation are potentially important targets for the treatment of T2D. Protein kinase D1 (PKD1) integrates diverse signals in the beta cell and plays a critical role in the control of insulin secretion. However, the role of beta-cell PKD1 in glucose homeostasis in vivo is essentially unknown. Using beta-cell specific, inducible PKD1 knock-out mice (βPKD1KO), we examined the role of beta-cell PKD1 under basal conditions and during high-fat feeding...
October 16, 2017: Diabetes
Ningbo Xia, Jichao Yang, Shu Ye, Lihong Zhang, Yanqin Zhou, Junlong Zhao, L David Sibley, Bang Shen
Glycolysis was thought to be the major pathway of energy supply in both fast replicating tachyzoites and slowly growing bradyzoites of Toxoplasma gondii. However its biological significance has not been clearly verified. The genome of T. gondii encodes two lactate dehydrogenases (LDH), which are differentially expressed in tachyzoites and bradyzoites. In this study, we knocked out the two LDH genes individually and in combination, and found that neither gene was required for tachyzoite growth in vitro under standard growth conditions...
October 13, 2017: Cellular Microbiology
Yue Cao, Dan Sun, Hao Ai, Hanyi Mei, Xue Liu, Shubin Sun, Guohua Xu, Yungen Liu, Yanshan Chen, Lena Q Ma
Arsenic (As) accumulation in rice grains poses health risk to humans. Plants including rice take up arsenate (AsV) by phosphate transporters. In this study, rice phosphate transporter OsPT4 (OsPht1;4) was investigated based on two independent T-DNA insertion mutants of OsPT4 (M1 and M2), which displayed stronger AsV resistance than wild types WT1 and WT2. When cultivated in medium (+P or -P) with AsV, ospt4 mutants accumulated 16-32% lower As in plants, suggesting that OsPT4 mediates AsV uptake. Analysis of the xylem sap showed that AsV concentrations in ospt4 mutants was 20-40% lower than WT controls under -P condition, indicating OsPT4 may also mediate AsV translocation...
November 7, 2017: Environmental Science & Technology
Vera Klenerova, Richard Kvetnansky, Sixtus Hynie
The activation of the HPA axis is the endocrine measure of stress responsiveness that is initiated by corticotropin-releasing hormone (CRH). CRH exerts its effects via CRHR1 and CRH-R2 receptors coupled to the cAMP signaling system and this process involves transcription factor cAMP-responsive element-binding protein (CREB).This study investigated the role of CRH and the possible involvement of CREB in gene regulation of CRH receptor, under basal conditions and after stress application in the pituitary. We used wild type (wt +/+) controls and CRH knock-out (CRH-KO -/-) male mice...
October 9, 2017: Cellular and Molecular Neurobiology
Paola Andrade, Daniel Caudepon, Teresa Altabella, Montserrat Arró, Albert Ferrer, David Manzano
Isoprenoids comprise the largest class of natural compounds and are found in all kinds of organisms. In plants, they participate in both primary and specialized metabolism, playing essential roles in nearly all aspects of growth and development. The enormous diversity of this family of compounds is extensively exploited for biotechnological and biomedical applications as biomaterials, biofuels or drugs. Despite their variety of structures, all isoprenoids derive from the common C5 precursor Isopentenil diphosphate (IPP)...
October 9, 2017: Plant Signaling & Behavior
Stéphane Peineau, Vincent Degos, Catherine Verney, Pierre Gressens
Proving endogenous GluN presence and functions in microglia require complementary steps to demonstrate (1) that GluN genes are transcripted and translated, (2) their cellular localization, (3) that the GluN are functional, and (4) the role of the functional GluN. The complete demonstration is performed by using mRNA detection technics, western blots, immunofluorescence, electrophysiology, calcium imaging, morphology studies, multiplex immunoassay together with conditional microglial Knock-Out mice and brain lesion models...
2017: Methods in Molecular Biology
Rolf Sprengel, Ahmed Eltokhi, Frank N Single
For the genetic alterations of NMDA receptor (NMDAR) properties like Ca(2+)-permeability or voltage-dependent gating in mice and for the experimental analysis of nonsense or missense mutations that were identified in human patients, single nucleotide mutations have to be introduced into the germ line of mice (Burnashev and Szepetowski, Curr Opin Pharmacol 20:73-82, 2015; Endele et al., Nat Genet 42:1021-1026, 2010). This can be done with very high precision by the well-established method of gene replacement, which makes use of homologous recombination in pluripotent embryonic stem (ES) cells of mice...
2017: Methods in Molecular Biology
Muna L Hilal, Maité M Moreau, Claudia Racca, Vera L Pinheiro, Nicolas H Piguel, Marie-Josée Santoni, Steve Dos Santos Carvalho, Jean-Michel Blanc, Yah-Se K Abada, Ronan Peyroutou, Chantal Medina, Hélène Doat, Thomas Papouin, Laurent Vuillard, Jean-Paul Borg, Rivka Rachel, Aude Panatier, Mireille Montcouquiol, Stéphane H R Oliet, Nathalie Sans
Planar cell polarity (PCP) signaling is well known to play a critical role during prenatal brain development; whether it plays specific roles at postnatal stages remains rather unknown. Here, we investigated the role of a key PCP-associated gene scrib in CA1 hippocampal structure and function at postnatal stages. We found that Scrib is required for learning and memory consolidation in the Morris water maze as well as synaptic maturation and NMDAR-dependent bidirectional plasticity. Furthermore, we unveiled a direct molecular interaction between Scrib and PP1/PP2A phosphatases whose levels were decreased in postsynaptic density of conditional knock-out mice...
December 1, 2017: Cerebral Cortex
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