• Molecules and Cells
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• v.44 no.5
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• pp.342-355
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• 2021

The microphthalmia-associated transcription factor family (MiT family) proteins are evolutionarily conserved transcription factors that perform many essential biological functions. In mammals, the MiT family consists of MITF (microphthalmia-associated transcription factor or melanocyte-inducing transcription factor), TFEB (transcription factor EB), TFE3 (transcription factor E3), and TFEC (transcription factor EC). These transcriptional factors belong to the basic helix-loop-helix-leucine zipper (bHLH-LZ) transcription factor family and bind the E-box DNA motifs in the promoter regions of target genes to enhance transcription. The best studied functions of MiT proteins include lysosome biogenesis and autophagy induction. In addition, they modulate cellular metabolism, mitochondria dynamics, and various stress responses. The control of nuclear localization via phosphorylation and dephosphorylation serves as the primary regulatory mechanism for MiT family proteins, and several kinases and phosphatases have been identified to directly determine the transcriptional activities of MiT proteins. In different immune cell types, each MiT family member is shown to play distinct or redundant roles and we expect that there is far more to learn about their functions and regulatory mechanisms in host defense and inflammatory responses.

• Molecules and Cells
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• v.46 no.12
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• pp.727-735
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• 2023

Stem cells require high amounts of energy to replicate their genome and organelles and differentiate into numerous cell types. Therefore, metabolic stress has a major impact on stem cell fate determination, including self-renewal, quiescence, and differentiation. Lysosomes are catabolic organelles that influence stem cell function and fate by regulating the degradation of intracellular components and maintaining cellular homeostasis in response to metabolic stress. Lysosomal functions altered by metabolic stress are tightly regulated by the transcription factor EB (TFEB) and TFE3, critical regulators of lysosomal gene expression. Therefore, understanding the regulatory mechanism of TFEB-mediated lysosomal function may provide some insight into stem cell fate determination under metabolic stress. In this review, we summarize the molecular mechanism of TFEB/TFE3 in modulating stem cell lysosomal function and then elucidate the role of TFEB/TFE3-mediated transcriptional activity in the determination of stem cell fate under metabolic stress.

• Proceedings of the KSAR Conference
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• 2003.06a
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• pp.76-76
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• 2003

Pluripotent embryonic stem cells can differentiate into beating cardiomyocytes with proper culture conditions and stimulants via embryo-like aggregates. We describe here the use of mouse embryonic stem (mES03) cells as a reproducible differentiation system for cardiomyocyte. mES03 cells growing in colonies were dissociated and allowed to re-aggregated in suspension [embryoid body (EB) formation〕. To induce cardiomyocytic differentiation, cells were exposed to 0.75% dimethyl sulfoxide (DMSO) during EB formation for 4 days and then another 4 days without DMSO (4+/4-). Thus treated EB was plated onto gelatin-coated dishes for differentiation. Spontaneously contracting colonies which appeared in approximately 4~5 days upon differentiation were mechanically dissected, enzymatically dispersed, plated onto coverslips, and then incubated for another 48~72 hrs. By RT-PCR, robust expression of cardiac myosin heavy chain $\alpha$, cardiac muscle heavy polypeptide 7 $\beta$($\beta$-MHC), cardiac transcription factor GATA4, and skeletal muscle-specific $\alpha$$_1$-subunit of the L-type calcium channel ($\alpha$$_1$CaC $h_{sm}$ ) were detected as early as 8 days after EB formation, but message of cardiac muscle-specific $\alpha$$_1$-subunit of the L-type calcium channel ($\alpha$$_1$CaCh) were reveled at a low level. In contrast, expression of myosin light chain (MLC-2V) and atrial natriuretic factor (ANF) were not detected during EB formation for 8 days. However, a strong expression of the atrial-specific ANF gene was expressed from day 8 onward, which were remained constant in EB. (cardiac specialization and terminal differentiation stage). Electrophysiological examination of spontaneously contracting cells showed ventricle-like action potential 17 days after the EB formation. This study indicates that mES03 cell-derived cardiomyocytes via 4+/4- protocol displayed biochemical and electrophysiological properties of subpopulation of cardiomyocytes.

• Korean Journal of Clinical Laboratory Science
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• v.51 no.2
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• pp.221-234
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• 2019

Cancer is genetically, metabolically and infectiously induced life threatening disorder showing aggressive growing pattern with invasive tendency. In order to prevent this global menace from jeopardizing human life, enormous studies on carcinogenesis and treatment for chemotherapy resistance have been intensively researched. Hinokitiol (${\beta}$-thujaplicin) extracted from heart wood of cupressaceous is a well-known bioactive compound demonstrating anti-inflammation, anti-bacteria and anti-cancer effects on several cancer types via apoptosis and autophagy. This study proposed that hinokitiol activates transcription factor EB (TFEB) nuclear translocation for autophagy and lysosomal biogenesis regardless of nutrient condition in cancer cells. Mitophagy and ${\beta}$-catenin translocation into the nucleus under treatment of hinokitiol on non-small cell lung cancer (NSCLC) cells and HeLa cells were investigated. Hinokitiol exerted cytotoxicity on HeLa and HCC827 cells; moreover, artificially induced autophagy by overexpression of TFEB granted imperfect sustainability onto HeLa cells. Taken together, hinokitiol is the prominent autophagy inducer and activator of TFEB nuclear translocation. Alternative cancer therapy via autophagy is pros and cons since the autophagy in cancer cells is related to prevention and survival mechanism depending on nutrition. To avoid paradox of autophagy in cancer therapy, fine-tuned regulation and application of hinokitiol in due course for successful suppressing cancer cells are recommended.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.100-100
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• 2003

Pluripotent embryonic stem (ES) cells differentiate spontaneously into beating cardiomyocytes via embryo-like aggregates. We describe the use of mouse embryonic stem (mES03) cells as a reproducible differentiation system for cardiomyocyte. To induce cardiomyocytic differentiation, mES03 cells were dissociated and allowed to aggregate (EB formation) at the presence of 0 75% dimethyl sulfoxide (DMSO) for 4 days and then another 4 days without DMSO (4+/4-). Thus treated EBs were plated onto gelatin-coated dish for differentiation. Spontaneously contracting colonies which appeared in approximately 4-5 days upon differentiation. Expression of cardiac-specific genes were determined by RT-PCR. Rebust expression of myosin light chain (MLC-2V), cardiac myosin heavy chain $\alpha$, cardiac muscle heavy polypeptide 7 $\beta(\beta$-MHC), cardiac transcription factor GATA4 and skeletal muscle-specific ${\alpha}_1$-subunit of the L-type calcium channel (${\alpha}_1 CaCh_{sm}$) were detected as early as 8 days after EB formation, but message of cardiac muscle-specific $\alpha$$_1$-subunit of the L-type calcium channel (${\alpha}_1$CaCh) were revealed at a low level. Strikingly, the expression of atrial natriuretic factor (ANF) was not detected. When spontaneous contracting cell masses were examined their electrophysiological features by patch-clamp technique, it showed ventricle-like action potential 17 days after the EB formation. This study indicates that mES03 cell-derived cardiomyocytes displayed biochemical and electrophysiological properties of cardiomyocytes and DMSO enhanced development of cardiomyocytes in 4+/4- method.

• The Korean Journal of Zoology
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• v.39 no.3
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• pp.239-247
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• 1996

An embryonic stem (ES) cell-based in vitro model system was examined to determine whether a simple differentiation of embryoid bodies (EB) in the suspension medium is useful to dissect early erythropoiesis. Characteristics of the differentiating EBs were monitored for their differentiation potential to generate hematopoietic cell types by general morphology, benzidine staining and two-step colony assays, and expressivity of several erythroid marker genes by the RT-PCR analysis for total cellular RNA prepared from the differentiating EBs. Every ematopoietic lineage cells were generated from the differentiating EBs with reproducible frequencies, similar to the other sophisticated differentiation protocols. Furthermore, the globin gene switching in differentiating ES cells paralleled the sequence of events found in the mouse embryo, and such that their expression was activated by at least 12 hrs later than those of erythroid-specific transcription factors, GATA-1 and Tal-1 The erythropoietic differentiation program initiated reproducibly and efficiently in this simple differentiation system in a suspension culture, such that this system may be useful for dissection of the molecular events of early erythropoiesis.

• IMMUNE NETWORK
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• v.24 no.3
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• pp.15.1-15.18
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• 2024

Osteoarthritis (OA) involves cartilage degeneration, thereby causing inflammation and pain. Cardiovascular diseases, such as dyslipidemia, are risk factors for OA; however, the mechanism is unclear. We investigated the effect of dyslipidemia on the development of OA. Treatment of cartilage cells with low-density lipoprotein (LDL) enhanced abnormal autophagy but suppressed normal autophagy and reduced the activity of transcription factor EB (TFEB), which is important for the function of lysosomes. Treatment of LDL-exposed chondrocytes with rapamycin, which activates TFEB, restored normal autophagy. Also, LDL enhanced the inflammatory death of chondrocytes, an effect reversed by rapamycin. In an animal model of hyperlipidemia-associated OA, dyslipidemia accelerated the development of OA, an effect reversed by treatment with a statin, an anti-dyslipidemia drug, or rapamycin, which activates TFEB. Dyslipidemia reduced the autophagic flux and induced necroptosis in the cartilage tissue of patients with OA. The levels of triglycerides, LDL, and total cholesterol were increased in patients with OA compared to those without OA. The C-reactive protein level of patients with dyslipidemia was higher than that of those without dyslipidemia after total knee replacement arthroplasty. In conclusion, oxidized LDL, an important risk factor of dyslipidemia, inhibited the activity of TFEB and reduced the autophagic flux, thereby inducing necroptosis in chondrocytes.