Phospholipase D and phosphatidylinositol-4-phosphate 5-kinase 1 are involved in the regulation of oligodendrocyte morphological differentiation

Kato, Yukino; Ochiai, Arisa; Seki, Yôtâro; Morimoto, Takako; Oizumi, Hiroaki; Ohbuchi, Katsuya; Mizoguchi, Kazuo; Yamamoto, Masahiro; Sakagami, Hiroyuki; Miyamoto, Yuki; Yamauchi, Junji

doi:10.1016/j.yexcr.2021.112654

Cited by 6 publications

(10 citation statements)

References 33 publications

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“…After a few days, cells grow to have a widespread membrane that is similar to the myelin membrane. Their cellular morphological phenotypes were identified as differentiated cells [ 27 ].…”

Section: Methodsmentioning

confidence: 99%

“…Cells were lysed in lysis buffer A (50 mM HEPES-NaOH, pH 7.5, 150 mM NaCl, 20 mM MgCl 2 , 1 mM phenylmethane sulfonyl fluoride, 1 microgram/mL leupeptin, 1 mM EDTA, 1 mM Na 3 VO 4 , 10 mM NaF, and 0.5% NP-40) [ 27 , 28 , 29 , 30 ]. After centrifugation, the supernatants were incubated with a non-denaturing sample buffer (also called a native polyacrylamide gel sample buffer; Nacalai Tesque) or a denaturing sample buffer (Nacalai Tesque) for non-denaturing and denaturing conditions, respectively.…”

Section: Methodsmentioning

confidence: 99%

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Hypomyelinating Leukodystrophy 8 (HLD8)-Associated Mutation of POLR3B Leads to Defective Oligodendroglial Morphological Differentiation Whose Effect Is Reversed by Ibuprofen

Sawaguchi

Suzuki

Oizumi

et al. 2022

Neurology International

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POLR3B and POLR3A are the major subunits of RNA polymerase III, which synthesizes non-coding RNAs such as tRNAs and rRNAs. Nucleotide mutations of the RNA polymerase 3 subunit b (polr3b) gene are responsible for hypomyelinating leukodystrophy 8 (HLD8), which is an autosomal recessive oligodendroglial cell disease. Despite the important association between POLR3B mutation and HLD8, it remains unclear how mutated POLR3B proteins cause oligodendroglial cell abnormalities. Herein, we show that a severe HLD8-associated nonsense mutation (Arg550-to-Ter (R550X)) primarily localizes POLR3B proteins as protein aggregates into lysosomes in the FBD-102b cell line as an oligodendroglial precursor cell model. Conversely, wild type POLR3B proteins were not localized in lysosomes. Additionally, the expression of proteins with the R550X mutation in cells decreased lysosome-related signaling through the mechanistic target of rapamycin (mTOR). Cells harboring the mutant constructs did not exhibit oligodendroglial cell differentiated phenotypes, which have widespread membranes that extend from their cell body. However, cells harboring the wild type constructs exhibited differentiated phenotypes. Ibuprofen, which is a non-steroidal anti-inflammatory drug (NSAID), improved the defects in their differentiation phenotypes and signaling through mTOR. These results indicate that the HLD8-associated POLR3B proteins with the R550X mutation are localized in lysosomes, decrease mTOR signaling, and inhibit oligodendroglial cell morphological differentiation, and ibuprofen improves these cellular pathological effects. These findings may reveal some of the molecular and cellular pathological mechanisms underlying HLD8 and their amelioration.

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“…After a few days, cells grow to have a widespread membrane that is similar to the myelin membrane. Their cellular morphological phenotypes were identified as differentiated cells [ 27 ].…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Hypomyelinating Leukodystrophy 8 (HLD8)-Associated Mutation of POLR3B Leads to Defective Oligodendroglial Morphological Differentiation Whose Effect Is Reversed by Ibuprofen

Sawaguchi

Suzuki

Oizumi

et al. 2022

Neurology International

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Section: Cell Culture and Differentiationmentioning

confidence: 99%

“…To induce differentiation, FBD-102b cells were cultured on tissue culture dishes coated with 100 µg/mL of polylysine in DMEM/F12 without FBS for 72 h in 5% CO 2 at 37 • C [15,16]. Cells bearing multiple processes (primary and secondary branches) from the cell bodies were identified as differentiated [15,16]. Differentiation was assessed in terms of three levels of completeness: Category 1 included cells with fewer than two primary branches; Category 2 included cells with fewer than three primary branches and without secondary branches (additional branches from the primary branches); and Category 3 included cells with more than three primary branches and with secondary branches as well as with widespread membranes.…”

Section: Cell Culture and Differentiationmentioning

confidence: 99%

“…Herein, we identify ACE2 itself as the negative regulator of cell morphological differentiation using FBD-102b cells, which are used as model cell lines undergoing differentiation of oligodendroglial cells [15,16]. RNA knockdown of ACE2 using the clustered regularly interspaced short palindromic repeats (CRISPR)-Cas system (CasRx-mediated guide RNA [gRNA]) [17,18] or its small interfering RNA (siRNA) resulted in promoting morphological differentiation.…”

Section: Introductionmentioning

confidence: 99%

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CRISPR/CasRx-Mediated RNA Knockdown Reveals That ACE2 Is Involved in the Regulation of Oligodendroglial Cell Morphological Differentiation

Kato

Tago

Fukatsu

et al. 2022

ncRNA

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Angiotensin-converting enzyme 2 (ACE2) plays a role in catalyzing angiotensin II conversion to angiotensin (1–7), which often counteracts the renin-angiotensin system. ACE2 is expressed not only in the cells of peripheral tissues such as the heart and kidney, but also in those of the central nervous system (CNS). Additionally, ACE2 acts as the receptor required for the entry of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), whose binding leads to endocytotic recycling and possible degradation of the ACE2 proteins themselves. One of the target cells for SARS-CoV-2 in the CNS is oligodendrocytes (oligodendroglial cells), which wrap neuronal axons with their differentiated plasma membranes called myelin membranes. Here, for the first time, we describe the role of ACE2 in FBD-102b cells, which are used as the differentiation models of oligodendroglial cells. Unexpectedly, RNA knockdown of ACE2 with CasRx-mediated gRNA or the cognate siRNA promoted oligodendroglial cell morphological differentiation with increased expression or phosphorylation levels of differentiation and/or myelin marker proteins, suggesting the negative role of ACE2 in morphological differentiation. Notably, ACE2′s intracellular region preferentially interacted with the active GTP-bound form of Ras. Thus, knockdown of ACE2 relatively increased GTP-bound Ras in an affinity-precipitation assay. Indeed, inhibition of Ras resulted in decreasing both morphological differentiation and expression or phosphorylation levels of marker proteins, confirming the positive role of Ras in differentiation. These results indicate the role of ACE2 itself as a negative regulator of oligodendroglial cell morphological differentiation, newly adding ACE2 to the list of regulators of oligodendroglial morphogenesis as well as of Ras-binding proteins. These findings might help us to understand why SARS-CoV-2 causes pathological effects in the CNS.

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miR-2013 negatively regulates phylogenetically conserved PIP5K involved in TLR4 mediated immune responses of amphioxus (Branchiostoma belcheri Tsingtaunese)

Cao

Fang

et al. 2022

Developmental & Comparative Immunology

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Phospholipase D and phosphatidylinositol-4-phosphate 5-kinase 1 are involved in the regulation of oligodendrocyte morphological differentiation

Cited by 6 publications

References 33 publications

Hypomyelinating Leukodystrophy 8 (HLD8)-Associated Mutation of POLR3B Leads to Defective Oligodendroglial Morphological Differentiation Whose Effect Is Reversed by Ibuprofen

Hypomyelinating Leukodystrophy 8 (HLD8)-Associated Mutation of POLR3B Leads to Defective Oligodendroglial Morphological Differentiation Whose Effect Is Reversed by Ibuprofen

CRISPR/CasRx-Mediated RNA Knockdown Reveals That ACE2 Is Involved in the Regulation of Oligodendroglial Cell Morphological Differentiation

miR-2013 negatively regulates phylogenetically conserved PIP5K involved in TLR4 mediated immune responses of amphioxus (Branchiostoma belcheri Tsingtaunese)

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