Noncanonical role of Golgi-associated macrophage TAZ in chronic inflammation and tumorigenesis

다국어 초록 (Multilingual Abstract)

Until now, Hippo pathway-mediated nucleocytoplasmic translocation has been considered the primary mechanism by which YAP and TAZ transcriptional coactivators regulate cell proliferation and differentiation via TEAD-mediated target gene expression. In this study, however, we found that TAZ, but not YAP, is associated with the Golgi apparatus in macrophages activated via Toll-like receptor (TLR) ligands during the resolution phase of inflammation. Golgi-associated TAZ enhanced vesicle trafficking and secretion of pro-inflammatory cytokines in M1 macrophage independent of the Hippo pathway. Depletion of TAZ in tumor-associated macrophages (TAM) promoted tumor growth by suppressing the recruitment of tumor-infiltrating lymphocytes (TIL). Moreover, in a diet-induced metabolic dysfunction-associated steatohepatitis (MASH) model, macrophage-specific deletion of TAZ ameliorated liver inflammation and hepatic fibrosis. Thus, targeted therapies being developed against YAP/TAZ-TEAD are ineffective in macrophages. Together, our results introduce Golgi-associate TAZ as a potential molecular target for therapeutic intervention to treat tumor progression and chronic inflammatory
diseases.

국문 초록 (Abstract)

지금까지 YAP 과 TAZ 전사 공동활성인자가 TEAD 매개 표적 유전자 발현을 통해 세포 증식과 분화를 조절하는 주요 메커니즘으로 Hippo 경로에 의한 핵과 세포질 간 이동이 알려져 있다. 그러나 이번 연구에서 우리는 염증 해결 단계에서 Toll-like receptor (TLR) 리간드에 의해 활성화된 대식세포에서 YAP이 아닌 TAZ가 골지체에 위치함을 발견했습니다. 골지와 연관된 TAZ는 Hippo 경로와 독립적으로 M1 대식세포에서 소포 수송과 염증성 사이토카인의 분비를 증진시켰습니다. 종양 연관 대식세포(TAM)에서 TAZ가 결핍되면 종양 침투 림프구(TIL)의 모집이 억제되어 종양 성장이 촉진되었습니다. 또한, 식이 유도 대사 기능 장애 관련 지방간염(MASH) 모델에서 대식세포 특이적 TAZ 결핍은 간 염증과 간 섬유화를 완화시켰습니다. 따라서 현재 임상에 사용중인 YAP/TAZ-TEAD를 표적으로 개발되고 있는 치료법은 대식세포에서 효과적이지 않다는 것을 밝혔습니다. 이러한 결과는 종양 진행과 만성 염증성 질환을 치료하기 위한 잠재적인 분자 표적으로 대식세포 특이적으로 골지와 연관된 TAZ를 제시했습니다.

목차 (Table of Contents)

• TABLE OF CONTENTS
• Table of Contents............................................................................ i
• List of Figures............................................................................... iv Abstract in English........................................................................... v
• Introduction................................................................................... 1 Chapter I. Discovering the Expression of TAZ in macrophages.................. 3 1. RESULTS..................................................................... 4 1.1 Induction of macrophage TAZ by TLR ligands...................... 4 1.2 TLR-mediated TAZ induction during the resolution phase of inflammation in macrophages............................................................... 5 1.3 Golgi localization of TAZ in TLR-stimulated macrophages....... 5 1.4 Hippo-independent regulation of TAZ in TLR-stimulated macrophages.................................................................. 6 1.5 Treatment with TEAD inhibitors in macrophages does not affect TAZ
• target genes.................................................................... 7
• Chapter II. Golgi-associated Macrophage TAZ plays a crucial role in chronic inflammation and tumorigenesis....................................................... 18
• 1. RESULTS.................................................................... 19 1.1 Golgi-associated TAZ regulates cytokine secretion M1-type macrophages................................................................. 19 1.2 Golgi-associated TAZ mediates broad protein trafficking...... 20 1.3 Induction of TAZ via LPS-activated TLR signaling in macrophages................................................................. 20 1.4 The role of macrophage TAZ in acute inflammatory responses..................................................................... 21 1.5 IL-6 signaling acts as upstream promoter of TAZ expression.... 22 1.6 Role of TAZ in tumor associated macrophages during immune cell
• i
• infiltration.................................................................... 23 1.7 Role of macrophage TAZ in inflammation and fibrosis of chronic liver diseases..................................................................... 24
• Materials and Methods.................................................................. 38
• 1.1 Public data analysis.................................................... 38
• 1.2 Mice..................................................................... 38
• 1.3 Cell culture.............................................................. 38
• 1.4 Isolation of bone marrow-derived macrophages (BMDMs)...... 39
• 1.5 Isolation of human monocyte from LRS chamber
• macrophage............................................................. 39
• 1.6 Co-culture of macrophage cells with cancer cells or normal cells...................................................................... 39
• 1.7 Chemical agents........................................................ 40
• 1.8 Bacterial strain and macrophage treatment ............................................................................. 40
• 1.9 Immunoblotting analysis.............................................. 40
• 1.10 Immunofluorescence microscopy.................................... 41 1.11 Retention Using Selective Hooks (RUSH) assay.................. 41 1.12 Immunoprecipitation assay.......................................... 42 1.13 Organelle fractionation................................................ 42 1.14 Antibodies............................................................... 42 1.15 RNA extraction, cDNA synthesis and quantitative real-time PCR (qRT- PCR)...................................................................... 43 1.16 siRNA transfection................................................... 45 1.17 RNA sequencing analysis............................................ 45 1.18 Chromatin immunoprecipitation (ChIP) assay and ChIP- seq........................................................................ 45 1.19 Quantification of inflammatory cytokines......................... 46 1.20 Animal experiments................................................... 47 1.21 Immunohistochemistry............................................... 48
• ii
• 1.22 Statistical analysis..................................................... 48
• Discussion................................................................................... 50 Reference.................................................................................... 55 Abstract in Korea .......................................................................... 61