Curbing Breast Cancer by Altering V-ATPase Action on F-Actin, Heterochromatin, ETV7 and mTORC2 Signaling.

IF 2.5 Q3 CELL BIOLOGY Cellular Physiology and Biochemistry Pub Date : 2024-06-12 DOI:10.33594/000000706
Zeina S Khan, Fazle Hussain
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引用次数: 0

Abstract

Background/aims: Motivated by the vacuolar proton pump's importance in cancer, we investigate the effects of proton pump inhibition on breast cancer cell migration and proliferation, F-actin polymerization, lamin A/C, heterochromatin, and ETV7 expressions, nuclear size and shape, and AKT/mTOR signaling.

Methods: Lowly metastatic MCF7 and highly metastatic MDA-MB-231 breast cancer cells were treated with 120 nM of proton pump inhibitor Bafilomycin A1 for 24 hours. Cell migration was studied with wound- scratch assays, ATP levels with a chemiluminescent assay; cell proliferation was quantified by a cell area expansion assay. Nuclear size and shape were determined using DAPI nuclear stain and fluorescence microscopy. The levels of F-actin, lamin A/C, heterochromatin, and ETV7 were quantified using both immunocytochemistry and western blots; p-mTORC1, p-mTORC2, mTOR, p-AKT, and AKT were measured by western blots.

Results: We reveal that proton pump inhibition reduces F-actin polymerization, cell migration, proliferation, and increases heterochromatin in both lowly and highly metastatic cells. Surprisingly, Bafilomycin decreases lamin A/C in both cell lines. Inhibition has different effects on ETV7 expression in lowly and highly metastatic cells, as well as nuclear area, perimeter, and circularity. Bafilomycin also significantly decreases p-mTORC1, p-MTORC2, and MTOR expression in both cell lines, whereas it significantly decreases p-AKT in lowly metastatic cells and surprisingly significantly increases p-AKT in highly metastatic cells. Our proton pump inhibition protocol reduces V-ATPase levels (~25%) within three hours. V-ATPase levels vary in time for both control and inhibited cells, and inhibition reduces cellular ATP.

Conclusion: Proton pumps promote F-actin polymerization and decrease heterochromatin, facilitating invasion. These pumps also upregulate both mTORC1 and mTORC2, thus highlighting the relevance of vacuolar proton pumps as metastatic cancer targets.

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通过改变 V-ATP 酶对 F-肌动蛋白、异染色质、ETV7 和 mTORC2 信号的作用抑制乳腺癌的发生
背景/目的:鉴于空泡质子泵在癌症中的重要性,我们研究了质子泵抑制剂对乳腺癌细胞迁移和增殖、F-肌动蛋白聚合、层粘连蛋白A/C、异染色质和ETV7表达、核大小和形状以及AKT/mTOR信号转导的影响:方法:用 120 nM 的质子泵抑制剂巴佛洛霉素 A1 处理低转移性 MCF7 和高转移性 MDA-MB-231 乳腺癌细胞 24 小时。细胞迁移采用伤口划痕试验进行研究,ATP 水平采用化学发光试验进行研究;细胞增殖采用细胞面积扩大试验进行量化。使用 DAPI 核染色和荧光显微镜测定细胞核的大小和形状。通过免疫细胞化学和 Western 印迹对 F-肌动蛋白、层粘连蛋白 A/C、异染色质和 ETV7 的水平进行了量化;通过 Western 印迹对 p-mTORC1、p-mTORC2、mTOR、p-AKT 和 AKT 进行了测定:结果:我们发现,质子泵抑制剂可减少低转移细胞和高转移细胞的 F-肌动蛋白聚合、细胞迁移和增殖,并增加异染色质。令人惊讶的是,巴佛洛霉素会降低两种细胞系中的层粘连蛋白 A/C。抑制作用对低转移细胞和高转移细胞中 ETV7 的表达以及核面积、周长和圆度都有不同的影响。巴佛洛霉素还能显著降低两种细胞系中的p-mTORC1、p-MTORC2和MTOR的表达,同时显著降低低度转移细胞中的p-AKT,而令人惊讶的是能显著增加高度转移细胞中的p-AKT。我们的质子泵抑制方案可在三小时内降低 V-ATPase 的水平(约 25%)。对照细胞和受抑制细胞的 V-ATP 酶水平随时间变化,抑制会降低细胞 ATP:结论:质子泵可促进 F-肌动蛋白聚合,减少异染色质,从而促进侵袭。这些泵还能上调 mTORC1 和 mTORC2,从而突出了空泡质子泵作为转移性癌症靶点的相关性。
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来源期刊
CiteScore
5.80
自引率
0.00%
发文量
86
审稿时长
1 months
期刊介绍: Cellular Physiology and Biochemistry is a multidisciplinary scientific forum dedicated to advancing the frontiers of basic cellular research. It addresses scientists from both the physiological and biochemical disciplines as well as related fields such as genetics, molecular biology, pathophysiology, pathobiochemistry and cellular toxicology & pharmacology. Original papers and reviews on the mechanisms of intracellular transmission, cellular metabolism, cell growth, differentiation and death, ion channels and carriers, and the maintenance, regulation and disturbances of cell volume are presented. Appearing monthly under peer review, Cellular Physiology and Biochemistry takes an active role in the concerted international effort to unravel the mechanisms of cellular function.
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