• 생명과학회지
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• 제34권5호
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• pp.333-338
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• 2024

키네신-1은 모터 도메인이 있는 두 개의 중쇄(KHC 또는 KIF5)와 모터 도메인이 없는 두 개의 경쇄(KLC)로 구성된 이형사량체 단백질이다. KIF5에는 KIF5A, KIF5B 및 KIF5C의 세 가지 subtype이 있으며, 카르복실(C)-말단 영역을 제외하고는 아미노산 상동성이 높다. KIF5A는 세포 내에서 화물을 운반하며, KIF5A의 C-말단 영역에 결합하는 매개 단백질은 키네신-1과 화물 사이를 연결한다. 키네신-1의 세포내 수송을 조절하는 단백질은 아직 충분히 확인되지 않았다. 본 연구는 리소좀의 세포 내 수송에 관여하는 ADP-ribosylation GTPase-activating protein 1 (ArfGAP1)과 KIF5A와의 결합을 확인하였다. KIF5A는 ArfGAP1의 C-말단 영역에 결합하고, ArfGAP1은 KIF5A의 C-말단 영역에 결합하지만 KIF5B, KIF5C, 키네신 경쇄 1 (KLC1) 또는 KIF3A와는 결합하지 않았다. ArfGAP 도메인을 가진 다른 동질형인 SMAP1과는 결합하지 않았다. 세포에서 KIF5A는 ArfGAP1과 같은 위치에서 발현하며, KIF5A, KIF5B 및 KLC1와 같이 면역 침전하였다. 그러나, KIF3B와는 같이 면역 침전하지 않았다. 이러한 결과들은 키네신-1은 세포내 화물 수송에서 ArfGAP1에 의해 조절될 수 있음을 시사한다.

• 생명과학회지
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• 제18권8호
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• pp.1059-1065
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• 2008

KIF21A는 kinesin superfamily에 속하는 분자 motor로서 미세소관을 따라서 분비소포를 운반한다. 최근의 연구결과 KIF21A 유전자 일부의 missense 돌연변이에 의하여 congenital fibrosis of the extraocular muscles (CFEOM) 1의 유발됨이 밝혀졌다. CFEOM1은 KIF21A의 돌연변이로 인하여 분화 발생과정에 occulo-motor신경과 neuromuscular junction 형성에 필요한 단백질을 이동시키지 못함으로써 유발된다. 본 연구에서는 효모 two-hybrid system을 사용하여 KIF21A의 WD-40 repeat domain과 결합하는 분자량이 작은 Purkinje cell protein-1 (Pcp-2), 또는 L7으로도 알려진 단백질을 분리하였다. Pcp-2는 효모 two-hybrid assay에서 KIF21A와 KIF21B의 WD-40 영역과는 결합하지만 다른 종류의 KIFs와는 결합하지 않았다. 또한 단백질간의 특이적 결합을 pull-down assay로 확인하였으며, 생쥐의 뇌 파쇄액에 Pcp-2 항체로 면역침강을 행하여 KIF21A를 확인한 결과 Pcp-2와 같이 침강하였다. 이러한 결과들은 KIF21A는 Pcp-2와 결합하며, 또한 Pcp-2는 KIF21A의 adaptor 단백질로서 세포 내 KIF21A의 수송에서 매개 단백질로 작용함을 시사한다.

• 생명과학회지
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• 제27권10호
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• pp.1191-1198
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• 2017

세포 내 소기관들과 소포들은 세포 내에서 미세소관을 따라 적절한 구획으로 수송된다. 이러한 세포 내 수송과정은 분자 모터단백질인 kinesin과 dynein에 의하여 이루어진다. Kinesin 1은 오징어 축삭돌기 세포질로부터 처음 분리되었으며 2개의 중쇄단위체(KHCs, 또는 KIF5s) 및 이와 결합하는 경쇄단위체(KLCs)의 복합체를 형성한다. KIF5s는C-말단 고리 영역을 통해 많은 다양한 단백질과 결합하는데, 아직 그 결합단백질들은 충분히 밝혀지지 않았다. 본 연구에서는 KIF5A 결합단백질을 분리하기 위하여 효모 two-hybrid 탐색을 수행하여 스트레스 과립형성과 mRNP 위치결정에 관여하는 Ras-GTPase-activating protein (GAP) Src homology3 (SH3)-domain-binding protein 2 (G3BP2)를 분리하였다. G3BP2는 KIF5A의 C-말단 고리 영역에 존재하는 73개 아미노산을 포함하는 영역과 결합하였다. 그러나 G3BP2는 KIF5B, KIF5C, KLC1, KIF3A와는 결합하지 않았다. KIF5A는 G3BP2의 arginine-glycine-glycine(RGG)/Gly-rich 도메인과 결합하지만 G3BP1과는 결합하지 않았다. HEK-293T세포에 G3BP2와 KIF5A를 발현하여 면역침강한 결과 G3BP2와 KIF5A는 같이 침강하였다. 또한 HEK-293T 세포 내의 전체에서 두 단백질은 같은 부위에 존재하였다. 이러한 결과들은 세포 내에서 G3BP2는 KIF5A와 결합하는 결합단백질로 확인 되었다.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2005년도 International Meeting of the Microbiological Society of Korea
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• pp.219.1-219
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• 2005

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2005년도 International Meeting of the Microbiological Society of Korea
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• pp.219.2-219
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• 2005

• 한국동물학회:학술대회논문집
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• 한국동물학회 2000년도 한국생물과학협회 학술발표대회
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• pp.256.1-256
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• 2000

No Abstract, See Full Text

• Journal of Korean Neurosurgical Society
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• 제67권3호
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• pp.364-375
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• 2024

Objective : Kinesin family member C1 (KIFC1), a non-essential kinesin-like motor protein, has been found to serve a crucial role in supernumerary centrosome clustering and the progression of several human cancer types. However, the role of KIFC1 in glioma has been rarely reported. Thus, the present study aimed to investigate the role of KIFC1 in glioma progression. Methods : Online bioinformatics analysis was performed to determine the association between KIFC1 expression and clinical outcomes in glioma. Immunohistochemical staining was conducted to analyze the expression levels of KIFC1 in glioma and normal brain tissues. Furthermore, KIFC1 expression was knocked in the glioma cell lines, U251 and U87MG, and the functional roles of KIFC1 in cell proliferation, invasion and migration were analyzed using cell multiplication, wound healing and Transwell invasion assays, respectively. The autophagic flux and expression levels matrix metalloproteinase-2 (MMP2) were also determined using imaging flow cytometry, western blotting and a gelation zymography assay. Results : The results revealed that KIFC1 expression levels were significantly upregulated in glioma tissues compared with normal brain tissues, and the expression levels were positively associated with tumor grade. Patients with glioma with low KIFC1 expression levels had a more favorable prognosis compared with patients with high KIFC1 expression levels. In vitro, KIFC1 knockdown not only inhibited the proliferation, migration and invasion of glioma cells, but also increased the autophagic flux and downregulated the expression levels of MMP2. Conclusion : Upregulation of KIFC1 expression may promote glioma progression and KIFC1 may serve as a potential prognostic biomarker and possible therapeutic target for glioma.

• BMB Reports
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• 제40권1호
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• pp.44-52
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• 2007

Kinesins, as a kind of microtubule-based motor proteins, have a conserved microtubule-binding site in their motor domain. Here we report that two homologous kinesins in Arabidopsis thaliana, KatB and KatC, contain a second microtubule-binding site in their tail domains. The prokaryotic-expressed N-terminal tail domain of the KatC heavy chain can bind to microtubules in an ATP-insensitive manner. To identify the precise region responsible for the binding, a serious of truncated KatC cDNAs encoding KatC N-terminal regions in different lengths, KatC1-128, KatC1-86, KatC1-73 and KatC1-63, fused to Histidine-tags, were expressed in E. coli and affinity-purified. Microtubule cosedimentation assays show that the site at amino acid residues 74-86 in KatC is important for microtubule-binding. By similarity, we obtained three different lengths of KatB N-terminal regions, KatB1-384, KatB1-77, and KatB1-63, and analyzed their microtubule-binding ability. Cosedimentation assays indicate that the KatB tail domain can also bind to microtubules at the same site as and in a similar manner to KatC. Fluorescence microscopic observations show that the microtubule-binding site at the tail domain of KatB or KatC can induce microtubules bundling only when the stalk domain is present. Through pull-down assays, we show that KatB1-385 and KatC1-394 are able to interact specifically with themselves and with each other in vitro. These findings are significant for identifying a previously uncharacterized microtubule-binding site in the two kinesin proteins, KatB and KatC, and the functional relations between them.

• Animal cells and systems
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• 제3권3호
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• pp.319-329
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• 1999

Onset of female puberty follows a series of prepubertal cellular and molecular events including changes of synaptic plasticity, synthetic and releasing activity and gene expression. Dramatic increase of gonadal steroid level is one of the most prominent changes before the onset of puberty. Based on the importance of steroid feedback upon the hypothalamus, we adopted an estrogen sterilized rat (ESR) model where 100 ng of 17$\eta$-estradiol were administered into neonatal pubs for 7 days after birth. To identify genes involved in the onset of female puberty, we applied PCR differential display using RNA samples derived from ESR and control rat hypothalami. About 100 out of more than 1000 RNA species examined displayed differential expression patterns between a 60-day old control rat and ESR. Sequence analysis of differentially amplified PCR products showed homology with genes such as mouse kinesin superfamily-associated protein 3 (KAP3) and several cDNAs previously described by others in mouse and human tissues. Several gene products such as 2-1 and 8-1 corresponded to novel DNA sequences. We analyzed mRNA levels of KAP3, 2-1 and 8-1 genes in the hypothalami derived from neonatal, 6-, 28-, 31-, and 40-day old rats. Northern blot analysis showed that mRNAs of KAP3, 2-1 and 8-1 genes were markedly increased before the initiation of puberty. Neonatal treatment of estrogen clearly inhibited prepubertal increases in KAP3, 2-1 and 8-1 mRNA levels. Therefore, these genes may play important roles in the initiation of hypothalamic puberty. In addition, intracerebroventricular (icv) injection of antisense KAP3 oligodeoxynucleotide (ODN) clearly delayed puberty initiation determined by vaginal opening, which further confirmed that KAP3 plays an important role in the regulation of puberty initiation.

• Journal of Ginseng Research
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• 제48권1호
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• pp.40-51
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• 2024

Background: Ginsenoside 20(S)-Rg3 shows promising tumor-suppressive effects in ovarian cancer via inhibiting NF-kB signaling. This study aimed to explore the downstream tumor suppressive mechanisms of ginsenoside Rg3 via this signaling pathway. Materials and methods: A systematical screening was applied to examine the expression profile of 41 kinesin family member genes in ovarian cancer. The regulatory effect of ginsenoside Rg3 on KIF20A expression was studied. In addition, we explored interacting proteins of KIF20A and their molecular regulations in ovarian cancer. RNA-seq data from The Cancer Genome Atlas (TCGA) was used for bioinformatic analysis. Epithelial ovarian cancer cell lines SKOV3 and A2780 were used as in vitro and in vivo cell models. Commercial human ovarian cancer tissue arrays were used for immunohistochemistry staining. Results: KIF20A is a biomarker of poor prognosis among the kinesin genes. It promotes ovarian cancer cell growth in vitro and in vivo. Ginsenoside Rg3 can suppress the transcription of KIF20A. GST pull-down and co-immunoprecipitation (IP) assays confirmed that KIF20A physically interacts with BTRC (β-TrCP1), a substrate recognition subunit for SCF^β-TrCP E3 ubiquitin ligase. In vitro ubiquitination and cycloheximide (CHX) chase assays showed that via interacting with BTRC, KIF20A reduces BTRC-mediated CDC25A poly-ubiquitination and enhances its stability. Ginsenoside Rg3 treatment partly abrogates KIF20A overexpression-induced CDC25A upregulation. Conclusion: This study revealed a novel anti-tumor mechanism of ginsenoside Rg3. It can inhibit KIF20A transcription and promote CDC25A proteasomal degradation in epithelial ovarian cancer.