• 생명과학회지
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• 제32권2호
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• pp.161-166
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• 2022

세포의 유사분열 과정에서 Spindle Assembly Checkpoint (SAC)는 키네토코어와 방추체의 미세소관의 연결을 확인하여 오류 없이 염색체 분열이 진행되도록 돕는 역할을 한다. SAC는 30년이 넘는 오랜 기간 동안 많은 연구자들에 의해 연구되었다. 하지만 단 하나의 연결되지 않은 키네토코어가 SAC를 어떻게 활성화시킬 수 있는지에 대해서는 그 기작이 명확히 밝혀지지 않았다. SAC의 핵심 단백질은 Mad1, Mad2. Mad3 (상위 진핵세포에서는 BubR1), Bub1, Bub3, Cdc20를 포함하는데, 이 단백질 모두 SAC의 활성화에 필요하다. SAC의 활성화에 핵심적인 단계는 미세소관과 연결되지 않은 키네토코어에서 Mad2과 Cdc20가 결합하여 복합체를 만드는 것인데, 이 과정은 화학반응에서 쉽게 일어나지 않는 반응이다. Mad2와 Cdc20가 어떻게 키네토코어로 갈 수 있는지에 대해서는 잘 알려져 있었지만, 어떻게 Mad2과 Cdc20가 결합하여 복합체 만들 수 있는지에 대해서는 알려지지 않았다. 최근 다른 실험 방법을 이용한 두 개의 다른 논문들이 어떻게 미세소관과 연결되지 않은 키네토코어에서 Mad2-Cdc20 복합체를 형성하는 지에 대한 핵심적인 기작을 밝혔다. 이 연구들은 단 하나의 연결되지 않은 키네토코어가 SAC 활성화시킬 수 있다는 것에 대한 가설을 뒷받침하고 있다. 본 논문에서는 SAC 활성화에 중요한 주요 기작들을 정리하고, SAC에 과한 최신 연구들을 자세히 살펴본 후, 이 결과들이 세포 분열 연구 분야에 있어서 어떻게 기여했는지 논의할 것이다.

• BMB Reports
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• 제39권6호
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• pp.709-716
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• 2006

Mammalian polo-like kinase 1 (Plk1) acts at various stages in early and late mitosis. Plk1 localizes at the centrosome and maintains this position through mitosis. Thereafter Plk1 moves to the kinetochore and midbody region, important sites during chromosome separation and cytokinesis. The catalytic domain of Plk1 is in the N-terminus region, whereas the non-catalytic region in the C-terminus of Plk1 has a conserved motif, named the Polobox. This motif is critical for Plk localization. EGFP proteins fused with the N-terminus and C-terminus of Plk1 localize in the nucleus and centrosomes, respectively. The core sequences of the polo-box (50 amino acids) also localize in Plk1 target organelles. To screen for domain-specific target proteins of Plk1, we constructed an N-terminal domain and a tandem repeat polo-box motif, and used them as templates in a yeast two-hybrid screen. The HeLa cell cDNA library indicated several proteins including the centrosome/kinetochore components or regulators, to be characterized as positive clones. Through in vitro protein binding analyses, we confirmed an interaction between these proteins and Plk1. The data reported from this study indicate that the N- and C- termini of Plk1 may function through recruitment and/or activation of domain-specific target proteins in dividing cells. Additionally, tandem repeats of the conserved core motif of the polo-box are sufficient for targeting and may be useful as a centrosome/kinetochore-specific targeting peptide.

• Molecules and Cells
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• 제27권6호
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• pp.697-701
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• 2009

We previously identified cancer-upregulated gene 2 (CUG2) as a commonly up-regulated gene in various human cancer tissues, especially in ovary, liver, and lung (Lee et al., 2007a). CUG2 was determined to be a nuclear protein that exhibited high proto-oncogenic activities when overexpressed in NIH3T3 mouse fibroblast cells. To identify other cellular functions of CUG2, we performed yeast two-hybrid screening and identified CENP-T, a component of CENP-A nucleosome complex in the centromere, as an interacting partner of CUG2. Moreover, CENP-A, the principle centromeric determinant, was also found in complex with CENP-T/CUG2. Immunofluorescent staining revealed the co-localization of CUG2 with human centromeric markers. Inhibition of CUG2 expression drastically affected cell viability by inducing aberrant cell division. We propose that CUG2 is a new component of the human centromeric complex that is required for proper chromosome segregation during mitosis.

• Journal of Applied Biological Chemistry
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• 제51권1호
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• pp.1-6
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• 2008

Estrogens, a group of steroid compounds functioning as the primary female sex hormone, play an important role in the development and progression of breast cancer. In this study, using a novel annealing control primer-based GeneFishing PCR technology, five differentially expressed genes (DEGs), expressed using 10nM mitogenic estrogens, $17{\beta}$-estradiol (E2) and $16{\alpha}$-hydroxyestrone ($16{\alpha}$-OHE1), were selected from the estrogen receptor (ER)-positive MCF-7 human breast cancer cells. The DEGs, MRPL42, TUBA1B, SSBP1, KNCT2, and RUVBL1, were identified by comparison with the known genes via direct sequencing and sequence homology search in BLAST. Quantitative real-time PCR data showed that two DEGs, tubulin ${\alpha}1b$ and kinetochore associated 2, were greater than 2-fold upregulated by E2 or $16{\alpha}$-OHE1. Both genes could be new biomarkers for the treatment and prognosis of cancers, and further study may provide insights into the molecular mechanisms underlying development and progression of breast cancer.

• Asian Pacific Journal of Cancer Prevention
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• 제15권20호
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• pp.8623-8629
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• 2014

Background: As an important component of the NDC80 kinetochore complex, NUF2 is essential for kinetochore-microtubule attachment and chromosome segregation. Previous studies also suggested its involvement in development of various kinds of human cancers, however, its expression and functions in human hepatocellular carcinoma (HCC) are still unclear. Materials and Methods: In the present study, we aimed to test the hypothesis that NUF2 is aberrant in human HCCs and associated with cell growth. Results: Our results showed significantly elevated expression of NUF2 in human HCC tissues compared to adjacent normal tissues, and high expression of NUF2 in HCC cell lines. Using lentivirus-mediated silencing of NUF2 in HepG2 human HCC cells, we found that NUF2 depletion markedly suppressed proliferation and colony formation capacity in vitro, and dramatically hampered tumor growth of xenografts in vivo. Moreover, NUF2 silencing could induce cell cycle arrest and trigger cell apoptosis. Additionally, altered levels of cell cycle and apoptosis related proteins including cyclin B1, Cdc25A, Cdc2, Bad and Bax were also observed. Conclusions: In conclusion, these results demonstrate that NUF2 plays a critical role in the regulation of HCC cell proliferation and apoptosis, indicating that NUF2 may serve as a potential molecular target for therapeutic approaches.

• 한국자기공명학회논문지
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• 제19권2호
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• pp.88-94
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• 2015

Bub1 is one of the spindle checkpoint proteins and plays a role in recruitment of the related proteins to kinetochore. Here, we studied the structural characteristic of the evolutionarily conserved 160 amino acid region in the N-terminus (hBub1 CD1), using Circular Dichroism (CD) and NMR. Our CD results showed that hBub1 CD1 is a highly helical protein and its structure was affected by pH: as pH was elevated to basic pH, the helical propensity increased. This could be related to the surface charge of the hBub1 CD1. However, the structural change did not largely depend on the salt concentration, though the thermal stability a little increased. The previous NMR analysis revealed that the hBub1 CD1 adopts eight helices, which is consistent with the CD result. Our result would be helpful for evaluating the molecular mechanism of the hBub1 CD1 and protein-protein interactions.

• Molecules and Cells
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• 제39권9호
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• pp.669-679
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• 2016

N-acetyl-D-glucosamine kinase (GlcNAc kinase or NAGK) primarily catalyzes phosphoryl transfer to GlcNAc during amino sugar metabolism. Recently, it was shown NAGK interacts with dynein light chain roadblock type 1 (DYNLRB1) and upregulates axo-dendritic growth, which is an enzyme activity-independent, non-canonical structural role. The authors examined the distributions of NAGK and NAGK-dynein complexes during the cell cycle in HEK293T cells. NAGK was expressed throughout different stages of cell division and immunocytochemistry (ICC) showed NAGK was localized at nuclear envelope, spindle microtubules (MTs), and kinetochores (KTs). A proximity ligation assay (PLA) for NAGK and DYNLRB1 revealed NAGK-dynein complex on nuclear envelopes in prophase cells and on chromosomes in metaphase cells. NAGK-DYNLRB1 PLA followed by Lis1/NudE1 immunostaining showed NAGK-dynein complexes were colocalized with Lis1 and NudE1 signals, and PLA for NAGK-Lis1 showed similar signal patterns, suggesting a functional link between NAGK and dynein-Lis1 complex. Subsequently, NAGK-dynein complexes were found in KTs and on nuclear membranes where KTs were marked with CENP-B ICC and nuclear membrane with lamin ICC. Furthermore, knockdown of NAGK by small hairpin (sh) RNA was found to delay cell division. These results indicate that the NAGK-dynein interaction with the involvements of Lis1 and NudE1 plays an important role in prophase nuclear envelope breakdown (NEB) and metaphase MT-KT attachment during eukaryotic cell division.

• Journal of Microbiology
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• 제39권4호
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• pp.286-292
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• 2001

During mitosis. the proper segregation of duplicated chromosomes is corrdinated by a spindle check-point. The bifurcated spindle checkpoint blocks cell cycle progression at metaphase by monitoring unattached kinetochores and inhibits mitotic exit in response to the misorientation of the mitotic spin- dle Ibd1p/Bfa1p is a spindle checkpoint regulator of budding yeast in the Bub2p checkpoint pathway for mitotic exit and its disruption abolishes mitotic arrest when proper organization of the mitotic spin-dls inhibited. Ibd1p/Bfa1p localizes to the spindle pole body, a microtublue-organizing center in yeast, and its overexpression arrests the cell cycle in 80% of cells with an enlarged budy at mitosis and in 20 % of cells with multiple buds. In this study, we found that the C-terminus of Ibd1p/Bfa1p phys-ically interacts with Skp1p, a key component of SCF (Skp1/cullin/F-box) complex for ubiquition-medi-ated proteolysis of cel cycle regulatores as well as an evolutionally conserved kinetochore protein for cell cycle progression. A putative F-box motif was found in the C-terminus of Ibd1p/Bfa1p and its function was investigated by making mutants of conserved residues in the motif. These Ibd1p/Bfa1p mutants of a putative F-box interacted with SKp1p in vitro by two-hybrid assays as wild type Ibd1p/Bfa1p. Also these Ibd1p/Bfa1p utants displayed the overexpression phenotypes of wild type Ibd1p, when over-expressed under inducible promoters . These results suggest that a putative F-box motif of Ibd1p/Bfa1p is not essential for the interaction with SKp1p and its function in mitotic exit and cytokinesis.

• 한국산림과학회지
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• 제77권3호
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• pp.338-350
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• 1988

남한(南韓)에 자생(自生) 또는 도입재배(導入栽培)되고 있는 향나무속(屬) 종류(種類)들의 분류학적(分類學的) 문제(問題)들을 해결(解決)하고 재정비(再整備)할 목적(目的)으로 자생종(自生種) 7종류(種類)와 조입종중(導入種中)에서 많이 재배(栽培)되고 유용(有用)하게 이용(利用)되고 있는 원예종(園藝種) 4종류(種類)를 재료(材料)로 하여 각종류(各種類)의 꽃의 구조(構造), 구과(毬果), 종자(種子), 가지습성(習性), 기본수형(基本樹形) 및 잎의 형태적(形態的) 특징(特徵)을 조사(調査)해서 속(屬)과 종(種)의 검사표(檢査表)를 작성(作成)했으며 외부형태적(外部形態的) 특징(特徵)으로는 종간(種間)의 식별(識別)이 모호(模糊)한 것을 명확(明確)히 구명(究明)하기 위(爲)해서 근단(根端)을 재료(材料)로 하여 염색체(染色體)의 기본수(基本數)와 핵형분석(核型分析)을 하고 그 결과(結果)로 각(各) 종간(種間)의 차이점(差異點)을 비교분석(比較分析)하였던바 전종(全種)의 염색체장(染色體長)은 $9.0{\sim}15{\mu}m$, 향나무, 심향나무, 가이즈가향나무, 섬향나무(일본산(日本産))은 4배체(倍體)(4n=44), 옥향나무, 눈향나무, 뚝향나무, 연필향나무, 노간주나무, 좀노간주나무, 해변노간주나무는 2배체(倍體)(2n=22)이고 향나무절(節)의 종(種)들은 염색체(染色體)길이 및 동원체(動原體)의 위치(位置)의 변이(變異)가 대단(大端)히 심(甚)하였다. 노간주나무절절(節)의 종(種)들은 11번 염색체(染色體)가 t형(型)이고, m형(型) 염색체(染色體)가 많은 비교적(比較的) 균일(均一)하고 단순(單純)한 염색체(染色體) set를 이루는 것이 향나무절(節)과 뚜렷이 다른 세포학적(細胞學的)인 특징(特徵)이다. 향나무절(節)에서는 대도시(大都市)에 여러 종류(種類)들이 혼식(混植)되어 잇는 곳에서는 동종(同種)의 지역품종(地域品種) ecotype간(間), 때로는 기본종(基本種)과 변종간(變種間)에 교배(交配)가 일어나고 있기 때문에 향나무 종내개체(種內個體)들 간(間)에는 형태적(形態的)으로 차이(差異)가 다양(多樣)하다. 상기(上記) 결과(結果)들을 종합(綜合)하여 향나무속(屬)의 특성(特性)을 세포학적(細胞學的)으로 명확(明確)히 식별(識別)하고 종간(種間)의 고유특성(固有特性)을 밝히고 종(種)의 특징(特徵)을 기재(記載)하였다.