Jihyun Lee

Heritable gene knockout in Caenorhabditis elegans by direct injection of Cas9-sgRNA ribonucleoproteins.

We present a novel method of targeted gene disruption that involves direct injection of recombinant Cas9 protein complexed with guide RNA into the gonad of the nematode Caenorhabditis elegans. Biallelic mutants were recovered among the F1 progeny, demonstrating the high efficiency of this method.

Hypoxia-inducible Factor-1 (HIF-1)-independent Hypoxia Response of the Small Heat Shock Protein hsp-16.1 Gene Regulated by Chromatin-remodeling Factors in the Nematode Caenorhabditis elegans

Background: To characterize an unknown molecular basis of the hif-1-independent hypoxia response, we used the nematode hsp-16.1 gene as a model. Results: HMG-1.2, together with chromatin-remodeling factors and calcium ions, is involved in the hypoxia response of hsp-16.1. Conclusion: Chromatin modification is important for the hypoxia response of hsp-16.1 in an HIF-1-independent manner. Significance: We report an alternative regulatory pathway for the HIF-1-independent hypoxia response. Oxygen deprivation is accompanied by the coordinated expression of numerous hypoxia-responsive genes, many of which are controlled by hypoxia-inducible factor-1 (HIF-1). However, the cellular response to hypoxia is not likely to be mediated by HIF-1 alone, and little is known about HIF-1-independent hypoxia responses. To better establish the molecular mechanisms of HIF-1-independent hypoxia responses, we sought to characterize the molecular basis of the hypoxia response of the hsp-16.1 gene in the nematode Caenorhabditis elegans; this gene has been shown to be induced by hypoxia independently of hif-1. Using affinity purification followed by LC-MS/MS, we identified HMG-1.2 as a protein that binds to a specific promoter region under hypoxic conditions. By systematic prediction followed by validation of these interactions through RNAi, we identified the chromatin modifiers isw-1 and hda-1, histone H4, and NURF-1 chromatin-remodeling factors as new components of the hif-1-independent hypoxia response. These data suggest that the modulation of nucleosome positioning at the hsp-16.1 promoter may be important for the hypoxia response. In addition, we found that calcineurin acts independently of hif-1 to modulate the cellular response to hypoxia and that calcium ions are necessary for the induction of hsp-16.1 under hypoxic conditions.

Efficient PRNP deletion in bovine genome using gene-editing technologies in bovine cells

abstract Even though prion (encoded by the PRNP gene) diseases like bovine spongiform encephalopathy (BSE) are fatal neurodegenerative diseases in cattle, their study via gene deletion has been limited due to the absence of cell lines or mutant models. In this study, we aim to develop an immortalized fibroblast cell line in which genome-engineering technology can be readily applied to create gene-modified clones for studies. To this end, this study is designed to 1) investigate the induction of primary fibroblasts to immortalization by introducing Bmi-1 and hTert genes; 2) investigate the disruption of the PRNP in those cells; and 3) evaluate the gene expression and embryonic development using knockout (KO) cell lines. Primary cells from a male neonate were immortalized with Bmi-1and hTert. Immortalized cells were cultured for more than 180 days without any changes in their doubling time and morphology. Furthermore, to knockout the PRNP gene, plasmids that encode transcription activator-like effector nuclease (TALEN) pairs were transfected into the cells, and transfected single cells were propagated. Mutated clonal cell lines were confirmed by T7 endonuclease I assay and sequencing. Four knockout cell lines were used for somatic cell nuclear transfer (SCNT), and the resulting embryos were developed to the blastocyst stage. The genes (CSNK2A1, FAM64A, MPG and PRND) were affected after PRNP disruption in immortalized cells. In conclusion, we established immortalized cattle fibroblasts using Bmi-1 and hTert genes, and used TALENs to knockout the PRNP gene in these immortalized cells. The efficient PRNP KO is expected to be a useful technology to develop our understanding of in vitro prion protein functions in cattle.

Identification of Genes Interacting with rnt-1 Through Large-Scale RNAi Screening in Caenorhabditis elegans

Although many critical roles of the RUNX family proteins have already been identified, little attention has been given to how these proteins interact with other factors. Elucidating RUNX protein interactions will help extend our understanding of their roles in normal development and tumorigenesis. In this study, we performed large-scale RNAi screening to identify genes that genetically interact with rnt-1, the sole homolog of RUNX protein in the nematode Caenorhabditis elegans. To this end, we took advantage of the fact that C. elegans can survive a severe loss of RNT-1 function with only mild phenotypes, and we looked for genes that caused a synthetic phenotype in the rnt-1 mutant background. We identified seven genes, three of which (cdk-8, cic-1, and sur-2) are involved in transcription, two of which (pgp-2 and cct-5) are involved in stress response, and two of which (D2045.7 and W09D10.4) are involved in signaling cascades, according to their functional gene ontology terms. We further confirmed that the CDK8-containing mediator complex genetically interacts with RNT-1 by showing that knockdown of each component of the CDK8 mediator complex caused a synthetic phenotype, that is, the exploded intestine through the vulva (Eiv) phenotype, in the rnt-1 mutant background. We also identified a putative target gene, acs-4, which is regulated by the RNT-1 and CDK8 mediator complex. Our results strengthen the notion that the CDK8 mediator complex may also act together with RUNX proteins in mammals.

Analysis of Studies on the Effect of Aquatic Exercise Program

Purpose: The purpose of this study was to analyze the effect of aquatic exercise and to identify directions for future research. Methods: Twenty one articles from 2 sites of article database (1990-2010) and 6 from Korean nursing journals were reviewed. Results: There were 20 quasi experimental design and 1 randomized controlled design. Major subjects of research were older women with arthritis. The most common type of exercise was Kim (1994)s form. The most frequent duration of exercise was over 6 weeks. Outcome measurement was classified to physical and psychosocial variables. For physical outcomes, pain release was found in 13 articles among 15 articles, muscle strength improvement in 5 articles among 10 articles, flexibility improvement in 5 articles among 9 articles, and balance improvement in 3 articles among 6 articles. For psychosocial outcomes, improvement of quality of life was found in 3 articles among 7 articles and self-efficacy improvement in 5 articles among 7 articles. Conclusion : The effect of aquatic exercise over 6 weeks improves the physical and psychosocial functions. It is suggested that meta analysis needs to be done to integrate the various results.

Expressive aphasia as the manifestation of hyperglycemic crisis in type 2 diabetes

Copyright

Nurses` Needs and Priority for Developing Advanced Nursing Education Program in Musculoskeletal Care

Purpose: The purpose of this study was to assess nurses` educational needs for developing advanced nursing education program for musculoskeletal care. Methods: The participants consisted of 272 clinical nurses who had experience of taking care of patients with musculoskeletal problems. A structured questionnaire with 28 items was used to assess educational needs and priority perceived by clinical nurses. Collected data were analyzed using SPSSWIN V.18.0 program for descriptive statistics. This survey was conducted in accordance with standard ethical guidelines for all research involving human participants. Results: While the majority (93.4%) perceived needs for advanced education program, about half of the participants (48.5%) reported no educational experience about musculoskeletal care. The clinical nurses perceived important for 23 out of 28 areas to be included in advanced nursing education. Especially, the areas of pain management, surgical care, and medication were perceived high priority for education program in caring musculoskeletal patients. Conclusion: The study findings showed that most nurses perceived the need for advanced nursing education program in musculoskeletal care, and the areas with higher priority as perceived by nurses should be reflected in the program.

A Study of Pain, Depression, and Adjustment to Military Life of Soldiers with Low Back Pain

Purpose: The purpose of this study was to examine levels of pain, depression, and adjustment to military life in soldier who experienced low back pain, in order to produce fundamental data for the development of health improvement programs to prevent and manage low back pain in soldiers with low back pain. Methods: Subjects of this study consisted of 317 soldiers who had low back pain. Study instruments were Visual analog scale (VAS), the Center for Epidemiological Studies-Depression Scale (CES-D), and adjustment to military life scale. Collected data were analyzed by the SPSS Win 14.0 program. Results: Mean score of pain was 4.16 points, depression was 8.58 points, and adjustment to military life was 69.15 points. The level of pain was negatively related to adjustment to military life (r