• Clinical and Experimental Reproductive Medicine
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• 제43권1호
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• pp.9-14
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• 2016

Objective: Autophagy contributes to the clearance and recycling of macromolecules and organelles in response to stress. We previously reported that vitrified mouse oocytes show acute increases in autophagy during warming. Herein, we investigate the potential role of Atg7 in oocyte vitrification by using an oocyte-specific deletion model of the Atg7 gene, a crucial upstream gene in the autophagic pathway. Methods: Oocyte-specific Atg7 deficient mice were generated by crossing Atg7 floxed mice and Zp3-Cre transgenic mice. The oocytes were vitrified-warmed and then subjected to in vitro fertilization and development. The rates of survival, fertilization, and development were assessed in the Atg7 deficient oocytes in comparison with the wildtype oocytes. Light chain 3 (LC3) immunofluorescence staining was performed to determine whether this method effectively evaluates the autophagy status of oocytes. Results: The survival rate of vitrified-warmed $Atg7^{f/f}$;Zp3-Cre ($Atg7^{d/d}$) metaphase II (MII) oocytes was not significantly different from that of the wildtype ($Atg7^{f/f}$) oocytes. Fertilization and development in the $Atg7^{d/d}$ oocytes were significantly lower than the $Atg7^{f/f}$ oocytes, comparable to the $Atg5^{d/d}$ oocytes previously described. Notably, the developmental rate improved slightly in vitrified-warmed $Atg7^{d/d}$ MII oocytes when compared to fresh $Atg7^{d/d}$ oocytes. LC3 immunofluorescence staining showed that this method can be reliably used to assess autophagic activation in oocytes. Conclusion: We confirmed that the LC3-positive signal is nearly absent in $Atg7^{d/d}$ oocytes. While autophagy is induced during the warming process after vitrification of MII oocytes, the Atg7 gene is not essential for survival of vitrified-warmed oocytes. Thus, induction of autophagy during warming of vitrified MII oocytes seems to be a natural response to manage cold or other cellular stresses.

• Journal of Microbiology and Biotechnology
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• 제20권12호
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• pp.1647-1652
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• 2010

Autophagy refers to the bulk degradation of cellular proteins and organelles through an autophagosome and plays a pivotal role in the development, cellular differentiation, aging, and elimination of aberrant structures. A failure of autophagy has been implicated in a growing list of mammalian disease states, including cancer and cardiomyopathy. Two ubiquitin-like systems are highly involved in autophagy, especially in the formation of autophagosomes. Here, we purified and characterized Atg7 (an E1-like enzyme), and Atg3 and Atg10 (E2-like enzymes) in order to gain an insight into the role played by ubiquitin-like systems in the formation of autophagosomes. Interestingly, we observed that Atg7 forms a homodimer to construct an active conformation, unlike other E1-like enzymes. Although Atg3 was detected as a monomer under physiological conditions, Atg10 existed in an oligomeric form, indicating that the mechanism by which Atg10 functions may differ from that of Atg3.

• Molecules and Cells
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• 제41권1호
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• pp.50-54
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• 2018

Atg5 and Atg7 have long been considered as essential molecules for autophagy. However, we found that cells lacking these molecules still form autophagic vacuoles and perform autophagic protein degradation when subjected to certain stressors. During this unconventional autophagy pathway, autophagosomes appeared to be generated in a Rab9-dependent manner by the fusion of vesicles derived from the trans-Golgi and late endosomes. Therefore, mammalian autophagy can occur via at least two different pathways; the Atg5/Atg7-dependent conventional pathway and an Atg5/Atg7-independent alternative pathway.

• Molecules and Cells
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• 제37권5호
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• pp.399-405
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• 2014

Autophagy targets cytoplasmic cargo to a lytic compartment for degradation. Autophagy-related (Atg) proteins, including the transmembrane protein Atg9, are involved in different steps of autophagy in yeast and mammalian cells. Functional classification of core Atg proteins in plants has not been clearly confirmed, partly because of the limited availability of reliable assays for monitoring autophagic flux. By using proUBQ10-GFP-ATG8a as an autophagic marker, we showed that autophagic flux is reduced but not completely compromised in Arabidopsis thaliana atg9 mutants. In contrast, we confirmed full inhibition of auto-phagic flux in atg7 and that the difference in autophagy was consistent with the differences in mutant phenotypes such as hypersensitivity to nutrient stress and selective autophagy. Autophagic flux is also reduced by an inhibitor of phosphatidylinositol kinase. Our data indicated that atg9 is phenotypically distinct from atg7 and atg2 in Arabidopsis, and we proposed that ATG9 and phosphatidylinositol kinase activity contribute to efficient autophagy in Arabidopsis.

• 한국산학기술학회논문지
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• 제12권8호
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• pp.3541-3546
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• 2011

자식작용은 난자 세포질의 단백질 고분자 물질과 세포 소기관 분해를 위해서 세포질 리소좀 통로에 유전적으로 작용하고 있으며 ATP합성과 단백질 재활용에 관여하고 있다. 이러한 자식작용은 난자 발달 과정에서 매우 중요하지만 세포질 내 자식작용의 동적 발달 과정의 근원적인 기전은 잘 알려지지 않고 있다. 따라서 본 연구에서는 초기 난자 발달 과정의 자식작용을 이해하기 위해서 쥐 난자 체외 성숙 과정에서 자식작용과 관련된 유전자들의 유전적 발현 수준을 분석하였다. Real Time RT-PCR 기법을 이용하여 유전자 Atg2a, Atg3, Atg4b, Atg5, Atg6, Atg7, Atg9a, 그리고 Wipi3 같은 모계에서 유전된 ATGs 군들의 유전자들은 수정난 유전체 활성화(ZGA) 이전 단계인 1세포기에서 높게 발현되었고, 그 후 이들 유전자들의 발현은 배반포 단계와 2세포기 4세포기 단계에서는 감소함을 알 수 있었다. Dram과 Atg9b 유전자들은 배반포와 1세포기 단계에서 발현됨으로서 모계 유전자이면서 ZGA에 의해서 발현되는 유전자임을 알 수 있었다. 한편 UIKI의 유전자 발현은 착상 전 단계에서 일정하게 나타남을 알 수 있었다. 하지만 Atg4d 유전자의 경우 4세포기에서부터 배 반포 단계까지 높게 나타남을 알 수 있었다. 이러한 결과로부터 생쥐 난자 발달 과정에서 자식작용과 관련된 유전자들은 초기 난자 발달과정에서 중요한 역할 과정임을 알 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제29권6호
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• pp.989-998
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• 2019

Autophagy is crucial for immune defense against Mycobacterium tuberculosis (Mtb) infection. Mtb can evade host immune attack and survival within macrophages by manipulating the autophagic process. MicroRNAs (miRNAs) are small, non-coding RNAs that are involved in regulating vital genes during Mtb infection. The precise role of miRNAs in autophagy with the exits of Mtb remains largely unknown. In this study, we found miR-1958, a new miRNA that could regulate autophagy by interacting with 3'UTR of autophagy-related gene 5 (Atg5). In addition, Mtb infection triggered miR-1958 expression in RAW264.7 cells. What's more, miR-1958 overexpression blocked autophagic flux by impairing the fusion of autophagosomes and lysosomes. Overexpression of miR-1958 reduced Atg5 expression and LC3 puncta while inhibition of miR-1958 brought an increase of Atg5 and LC3 puncta; the opposite results were observed in detection of p62. The survival of Mtb in RAW264.7 cells transfected with mimic of miR-1958 was enhanced. Taken together, our research demonstrated that a novel miR-1958 could inhibit autophagy by interacting with Atg5 and favored intracellular Mtb survival in RAW264.7 cells.

• Clinical and Experimental Reproductive Medicine
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• 제50권3호
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• pp.170-176
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• 2023

Objective: Autophagy is highly active in ovariectomized mice experiencing hormone deprivation, especially in the uterine mesenchyme. Autophagy is responsible for the turnover of vasoactive factors in the uterus, which was demonstrated in anti-Müllerian hormone receptor type 2 receptor (Amhr2)-Cre-driven autophagy-related gene 7 (Atg7) knockout (Amhr-Cre/Atg7^f/f mice). In that study, we uncovered a striking difference in the amount of sequestosome 1 (SQSTM1) accumulation between virgin mice and breeder mice with the same genotype. Herein, we aimed to determine whether repeated breeding changed the composition of mesenchymal cell populations in the uterine stroma. Methods: All female mice used in this study were of the same genotype. Atg7 was deleted by Amhr2 promoter-driven Cre recombinase in the uterine stroma and myometrium, except for a triangular stromal region on the mesometrial side. Amhr-Cre/Atg7^f/f female mice were divided into two groups: virgin mice with no mating history and aged between 11 and 12 months, and breeder mice with at least 6-month breeding cycles with multiple pregnancies and aged around 12 months. The uteri were used for Western blotting and immunofluorescence staining. Results: SQSTM1 accumulation, representing Atg7 deletion and halted autophagy, was much higher in virgin mice than in breeders. Breeders showed reduced accumulation of several vasoconstrictive factors, which are potential autophagy targets, in the uterus, suggesting that the uterine stroma was repopulated with autophagy-intact cells during repeated pregnancies. Conclusion: Multiple pregnancies seem to have improved the uterine environment by replacing autophagy-deficient cells with autophagy-intact cells, providing evidence of cell mixing.

• 생명과학회지
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• 제24권11호
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• pp.1180-1186
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• 2014

자가포식은 자가포식소체에서 리소좀으로 이어지는 경로를 통해 세포내에서 일어나는 자가분해 시스템으로 다양한 질병과 관련된 운동의 효과를 설명하는데 있어 중요한 역할을 하는 것으로 알려져 있다. 하지만 현재까지 보고된 대부분의 연구는 일회성 운동 혹은 저항성 운동에 의한 자가포식만을 조사한 것으로 일회성 운동과 만성적 운동을 함께 조사한 연구는 전무하다. 따라서 본 연구의 목적은 일회성 및 만성적 유산소운동이 자가포식에 미치는 영향을 알아보는 것이다. White-eyed mutant Drosophila melanogaster를 대상으로 하여 이들을 통제집단, 일회성 운동 집단, 일주일간 하루 2시간 또는 3시간씩 운동을 실시한 집단에 무선배정 하였고, 운동은 일회성 운동과 만성적 운동으로 구성하여 실시하였다. 정량 실시간 연쇄반응을 통해 Atg1, Atg6, Atg8a 유전자의 발현을 관찰한 결과 일회성 운동은 Atg8a mRNA 발현을 유의하게 증가시켰다. 그러나 Atg1, Atg6 mRNA 발현은 일회성 운동으로 유도되지 않았다. 또한 2시간의 만성적 운동에 의해서 Atg6 mRNA 발현이 유의하게 증가되었으나 Atg1, Atg8a mRNA 발현에는 유의한 변화가 나타나지 않았다. 반면 3시간의 만성적 운동을 실시한 결과 Atg1, Atg6, Atg8a mRNA 발현이 각각 57%, 37%, 71%로 모두 유의하게 증가된 것을 관찰할 수 있었다. 본 연구의 결과를 통해서 일회성의 운동은 선별된 자가포식 관련 유전자를 모두 활성화시키지는 못하지만 선택적으로 유도한다는 사실을 확인할 수 있었고, 만성적 운동은 자가포식 관련 유전자의 발현을 증가시킨다는 사실을 확인할 수 있었다. 따라서 본 연구는 일회성 및 만성적 유산소운동이 자가포식에 미치는 영향을 규명하였으며, 자가포식 관련 유전자가 일회성 운동과 만성적 운동에 의해 차별적으로 증가한다는 사실을 확인하였다는데 의의가 있다. 향후 운동에 의한 자가포식 연구에 있어 운동 형태 및 지속시간 등을 고려한 보다 세부적이고 체계적인 연구가 수행될 필요가 있을 것이라 사료되는 바이다.

• 한국세라믹학회지
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• 제35권10호
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• pp.1061-1069
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• 1998

The thermal instability of Al2TiO5 Ceramics was contrlled by solid solution with MgO SiO2 and ZrO2 through electrofusion in an arc furnace. The thermal expansion properties of Al2TiO5 composites show the hysteresis due to the strong anisotropy of The crystal axes of these material. These phenomena are ex-plained by the opening and closing of microcracks. The difference in microcracking temperatures e.g 587.6(ATG2), 405.9(ATG3) and 519.7$^{\circ}C$(ATG4) is caused by the difference in grain size and stabilizer type. The thermal shock behaviour under cyclic conditions between 750-1400-75$0^{\circ}C$ show no change in mi-crostructure and phase assemblage for all three stabilized specimens. After the thermal loading test at 110$0^{\circ}C$ for 100hrs. ATG1 and ATG2 materials decomposes completely to its components corundum and ru-tile in both cases. However with approximatelly 20% retention of the Al2TiO5 Thus in order to prevent decomposition of the stabilized material in the critical temperature range 800-130$0^{\circ}C$ it must be traversed within a short period of time.

• Clinical and Experimental Pediatrics
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• 제58권7호
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• pp.267-269
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• 2015

Antithymocyte globulin (ATG) is used as an immunosuppressive treatment (IST) to deplete clonal suppressor T cells in patients with severe aplastic anemia (SAA). The depletion of suppressor T cells by ATG may affect the activation of B cells, which results in an increased risk for autoimmune conditions. A 12-year-old boy was diagnosed with idiopathic SAA. As he did not have an human leukocyte antigen-matched sibling, he was treated with rabbit ATG (3.5 mg/kg/day for 5 days) and cyclosporine. Five months later, he became transfusion independent. However, 23 months after IST, he complained of mild hand tremors, sweating, weight loss, palpitations, and goiter. Results of thyroid function tests revealed hyperthyroidism (free thyroxine, 3.42 ng/dL; thyroid stimulating hormone [TSH], <0.01 nIU/mL; triiodothyronine, 3.99 ng/mL). Results of tests for autoantibodies were positive for the antimicrosome antibody and TSH-binding inhibitory immunoglobulin, but negative for the antithyroglobulin antibody and antinuclear antibody. He was treated with methimazole, and his symptoms improved. The patient has been disease free for 39 months after IST and 9 months after methimazole treatment. This case report suggests that although rare, rabbit ATG may have implications in the pathogenesis of autoimmune hyperthyroidism. Our findings suggest that thyroid function tests should be incorporated in the routine follow-up of SAA patients treated with ATG.