• Korean Journal of Microbiology
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• v.30 no.1
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• pp.37-41
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• 1992

The KEM1 gene is known to affect microtubule and spindle pole body function during the cell division cycle in Saccharomjyces cerevisiae. To identify new genes with functions similar or related to those of KEM1, we isolated a high copy suppressor gene (ROK1) that suppresses the kem1 mutation when cloned on a high copy number plasmid but not on a low copy number plasmid. Two clones which suppress both the benomyl hypersensitivity and the $Kar^{-}$ enhancing phenotype of kem1 null mutation were isolated and were shown to have a 9.0 kb identical insert by restriction endonuclease analysis. The restriction map constructed indicates that this suppressor gene, ROK1 is not KEM1. Subcloning experiments suggest that the functional region of ROK1 is at least 3.0kb in size.

• Korean Journal of Animal Reproduction
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• v.26 no.4
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• pp.361-368
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• 2002

The onset of pronucleus formation and DNA synthesis in porcine oocytes following the injection of porcine or murine sperm was determined in order to obtain insights into species-specific paternal factors that contribute to fertilization. After 44h in vitro maturation, spermatozoa was injected into the cytoplasm of oocytes. After injection, all oocytes were transferred to NCSU23 medium and cultured at 39'E under 5% CO2 in air. Similar frequencies of oocytes with female pronuclei were observed after injection with porcine sperm or with murine sperm. In contrast, male pronuclei formed 8 to 9 h following the injection of porcine sperm, and 6 to 8 h following the injection of murine sperm. After pronucleus formation maternally derived microtubules were assembled and appeared to move both male and female pronuclei to the oocyte center. A few porcine oocytes entered metaphase 22 h after the injection of murine sperm, but normal cell division was not observed. The mean time of onset of S-phase in male pronuclei was 9.7 h following porcine sperm injection and 7.4 h following mouse sperm injection. These results suggested that DNA synthesis was delayed in both pronuclei until the sperm chromatin fully decondensed, and the sperm nuclear decondensing activity and microtubule nucleation abilities of the male centrosome are cell cycle dependent.

• Korean Journal of Animal Reproduction
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• v.22 no.2
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• pp.195-202
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• 1998

We demonstrated, for the first time, pronuclear formation and apposition in porcine ooc-ytes following intracytoplasmic injection of porcine, human, bovine and mouse spermatozoon. Microtubule organization and chromatin configuration were investigated in these oocytes during pronuclear apposition. Following intracytoplasmic injection of porcine spermatozoon, the microtubular aster was organized from the neck of spermatozoon, and filled the whole cytoplasm. This male derived microtubules appear to move both pronuclei to the center of oocytes. In contrast, following injection of spermatozoa from different species such as human, bovine and mouse, microtubules were organized from the cortex of the oocytes and concentrated to the pronuclei, which seems to move both male and female pronuclei to the center of oocyte. This organization is similar to what has been shown in the parthenogenetically activated por-cine oocytes. These results suggested that the porcine, human, bovine and mouse sperm chromatin can be formed pronucleus and apposited in the center of oocytes in the absence of male derived microtubule when they were injected into porcine oocytes.

• Applied Microscopy
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• v.22 no.2
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• pp.46-65
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• 1992

In this study, we have made morphological and cytochemical observations to investigate the type of Golgi apparatus around the bile canaliculus. The animal (Wister, $220{\sim}250gm$) were divided into 4 groups; normal, hydrochol, colchicine and hydrochol-colchicine. The Golgi apparatus is classified into 16 different types from 4 different groups. In the normal group, we could observe 12 different types of the sixteen. Type I which showed convexed cisterns facing the bile canaliculi was most abundant of the types. In the hydrochol group, 14 types were observed. Type VII and type I showed convexed cisterns facing the bile canaliculus and were abundant. In the colchicine group, 11 different types were viewed and type XIV which showed intensely dilated cisterns without the polarity was predominant. In the hydrochol-colchicine group, we observed 3 different types. Type XIV clearly showed the highest percentage, although that type was less numerous in this group than in the colchicine group. In the hydrochol group, the Golgi apparatus showed a tendency to increase in numbers, while in the hydrochol-colchicine group the Golgi apparatus showed a tendency to decrease in numbers. The reactive products of thiamine pyrophosphatase and acid phosphatase were apparent over the distal Golgi cistern in the normal and hydrochol groups, but were decreased or not observed in the colchicine and hydrochol-colchicine groups. From the results, it is assumed that with the presence of the microtubule, Golgi cisterns are dilated with polarity after stimulation of secretion. Without the microtubule, the cistern becomes more intensely dilated and none polaric. Also the enzymes within the cisternal membrane become decreaed or absent and the Golgi apparatus decreases in numbers after activation of secretion.

• Molecules and Cells
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• v.22 no.2
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• pp.146-153
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• 2006

Cell polarity is critical for the division, differentiation, migration, and signaling of eukaryotic cells. RAX2 of budding yeast encodes a membrane protein localized at the cell cortex that helps maintain the polarity of the bipolar pattern. Here, we designate SPAC6f6.06c as $rax2^+$ of Schizosaccharomyces pombe, based on its sequence homology with RAX2, and examine its function in cell polarity. S. pombe $rax2^+$ is not essential, but ${\Delta}rax2$ cells are slightly smaller and grow slower than wild type cells. During vegetative growth or arrest at G1 by mutation of cdc10, deletion of $rax2^+$ increases the number of cells failing old end growth just after division. In addition, this failure of old end growth is dramatically increased in ${\Delta}tea1{\Delta}rax2$, pointing to genetic interaction of $rax2^+$ with $tea1^+$. ${\Delta}rax2$ cells contain normal actin and microtubule cytoskeletons, but lack actin cables, and the polarity factor for3p is not properly localized at the growing tip. In ${\Delta}rax2$ cells, and endogenous rax2p is localized at the cell cortex of growing cell tips in an actin- and microtubule-dependent manner. However, ${\Delta}rax2$ cells show no defects in cell polarity during shmoo formation and conjugation. Taken together, these observations suggest that rax2p controls the cell polarity of fission yeast during vegetative growth by regulating for3p localization.

• Animal cells and systems
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• v.12 no.2
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• pp.69-75
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• 2008

The tumor suppressor and transcription factor p53 is a key modulator of cellular stress responses, and can trigger apoptosis in many cell types including neurons. In this study, we have shown that Microtubule-associated protein 1B(MAP1B) light chain interacts with tumor suppressor p53. MAP1B is one of the major cytoskeletal proteins in the developing nervous system and essential in forming axons during elongation. We also demonstrate that both p53 and MAP1B-LC1 interact in the nucleus in HEK 293 cells. Indeed, we show that the MAP1B-LC1 negatively regulates p53-dependent transcriptional activity of a reporter containing the p21 promoter. Consequently, MAP1B light chain binds with p53 and their interaction leads to the inhibition of doxorubicin-induced apoptosis in HEK 293 cells. Furthermore, these examinations might be taken into consideration when knock-down of MAP1B-LC1 is used as a cancer therapeutic strategy to enhance p53's apoptotic activity in chemotherapy.

• Korean Journal of Microbiology
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• v.36 no.4
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• pp.306-311
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• 2000

Infection of Vero cells with herpes simplex virus type-1 (HSV-1) resulted in a series of changes in intra-cellular free calcium concentration $([Ca^{2+}]_i)$. A significant and maximal decrease $[Ca^{2+}]_i$ was observed at 4 hours postinfection (hr p.i.) in HSV-1-infected in Vero cells. Inactivation of HSV-1 with UV irradiation and heat treatment abolished HSV-1-induced decrease in $[Ca^{2+}]_i$ at 4 hr p.i. in Vero cells. And the degree of the decrease in $[Ca^{2+}]_i$ was dependent on the amount of input virus. Taxol, which stabilizes the polymerization of microtubule blocked HSV-1-induced decrease in $[Ca^{2+}]_i$ at 4 hr p.i., suggesting that microtubule may mediate the transport of HSV-1 nucleocapsid to the nucleus of infected cell. Treatment of HSV-1-infected Vero cells with metabolic inhibitors such as cycloheximide, cordycepin, or acyclovir partially reversed the decrease in $[Ca^{2+}]_i$ at 4 hr p.i.. Thus, it is suggested that HSV-1 induced decrease in $[Ca^{2+}]_i$ at 4 hr p.i. in Vero cells may play an important role in the multiplication of HSV-1.

• Physical Therapy Korea
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• v.14 no.2
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• pp.11-20
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• 2007

The development of neonatal neuromuscular system is accomplished by the functional interaction between the spinal neurons and its target cells, skeletal muscle cells, and the intrinsic and extrinsic factors affecting this process. The aim of this study was to identify the effect of suspension unloading (SU) and neuromuscular electrical stimulation (NMES) upon the development of the neonatal spinal cord. For this study, the neonatal rats were randomly divided into three groups: a control group, an experimental group I, and an experimental group II. The SU for experimental group I and II was applied from postnatal day (PD) 5 to PD 30, and the NMES for experimental group II was applied from PD 16 to PD 30 using NMES that gave isometric contraction with 10 Hz for 30 minutes twice a day. In order to observe the effect of SU and NMES, this study observed neutrophin-3 (NT-3) and microtubule associated protein 2 (MAP2) immunoreactivity in the lumbar spinal cord (L4-5) at the PD 15 and PD 30. The results are as follows. At PD 15, lumbar spinal cord of experimental group I and II had significantly lower NT-3 and MAP2 immunoreactivity than control group. It proved that a microgravity condition restricted the spinal development. At PD 30, lumbar spinal cord of control group and experimental group II had significantly higher NT-3 and MAP2 immunoreactivity than experimental group I. It proved that the NMES facilitated the spinal development by spinal cord-skeletal muscle interaction. These results suggest that weight bearing during the neonatal developmental period is essential for the development of neuromuscular development. Also, the NMES on its target skeletal muscle can encourage the development of the spinal cord system with a full supplementation of the effect of weight bearing, which is an essential factor in neonatal developmental process.

• BMB Reports
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• v.48 no.10
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• pp.583-588
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• 2015

Microtubule actin crosslinking factor 1 (MACF1), a widely expressed cytoskeletal linker, plays important roles in various cells by regulating cytoskeleton dynamics. However, its role in osteoblastic cells is not well understood. Based on our previous findings that the association of MACF1 with F-actin and microtubules in osteoblast-like cells was altered under magnetic force conditions, here, by adopting a stable MACF1-knockdown MC3T3-E1 osteoblastic cell line, we found that MACF1 knockdown induced large cells with a binuclear/multinuclear structure. Further, immunofluorescence staining showed disorganization of F-actin and microtubules in MACF1-knockdown cells. Cell counting revealed significant decrease of cell proliferation and cell cycle analysis showed an S phase cell cycle arrest in MACF1-knockdown cells. Moreover and interestingly, MACF1 knockdown showed a potential effect on cellular MTT reduction activity and mitochondrial content, suggesting an impact on cellular metabolic activity. These results together indicate an important role of MACF1 in regulating osteoblastic cell morphology and function.

• Molecules and Cells
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• v.38 no.10
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• pp.876-885
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• 2015

N-acetyl-D-glucosamine kinase (NAGK) plays an enzyme activity-independent, non-canonical role in the dendritogenesis of hippocampal neurons in culture. In this study, we investigated its role in axonal development. We found NAGK was distributed throughout neurons until developmental stage 3 (axonal outgrowth), and that its axonal expression remarkably decreased during stage 4 (dendritic outgrowth) and became negligible in stage 5 (mature). Immunocytochemistry (ICC) showed colocalization of NAGK with tubulin in hippocampal neurons and with Golgi in somata, dendrites, and nascent axons. A proximity ligation assay (PLA) for NAGK and Golgi marker protein followed by ICC for tubulin or dynein light chain roadblock type 1 (DYNLRB1) in stage 3 neurons showed NAGK-Golgi complex colocalized with DYNLRB1 at the tips of microtubule (MT) fibers in axonal growth cones and in somatodendritic areas. PLAs for NAGK-dynein combined with tubulin or Golgi ICC showed similar signal patterns, indicating a three way interaction between NAGK, dynein, and Golgi in growing axons. In addition, overexpression of the NAGK gene and of kinase mutant NAGK genes increased axonal lengths, and knockdown of NAGK by small hairpin (sh) RNA reduced axonal lengths; suggesting a structural role for NAGK in axonal growth. Finally, transfection of 'DYNLRB1 (74-96)', a small peptide derived from DYNLRB1's C-terminal, which binds with NAGK, resulted in neurons with shorter axons in culture. The authors suggest a NAGK-dynein-Golgi tripartite interaction in growing axons is instrumental during early axonal development.