Mertk deficiency affects macrophage directional migration via disruption of cytoskeletal organization.

Yong Tang,Qian Liu,Jiayi Xie,Dong Han,Qingjun Lu,Shen Wu,Jingxue Zhang,Qingxian Lu,Alissa M Weaver

doi:10.1371/journal.pone.0117787

Abstract

Mertk belongs to the Tyro3, Axl and Mertk (TAM) family of receptor tyrosine kinases, and plays a pivotal role in regulation of cytoskeletal rearrangement during phagocytosis. Phagocytosis by either professional or non-professional phagocytes is impaired in the Mertk deficient individual. In the present study, we further investigated the effects of Mertk mutation on peritoneal macrophage morphology, attachment, spreading and movement. Mertk-mutated macrophages exhibited decreased attachment, weak spreading, loss of spindle-like body shape and lack of clear leading and trailing edges within the first few hours of culture, as observed by environmental scanning electron microscopy. Time-lapse video photography recording showed that macrophage without Mertk conducted mainly random movement with oscillating swing around the cell body, and lost the directional migration action seen on the WT cells. Western blotting showed a decreased phosphorylation of focal adhesion kinase (FAK). Immunocytochemistry revealed that actin filaments and dynamic protein myosin II failed to concentrate in the leading edge of migrating cells. Microtubules were localized mainly in one side of mutant cell body, with no clear MTOC and associated radially-distributed microtubule bundles, which were clearly evident in the WT cells. Our results suggest that Mertk deficiency affects not only phagocytosis but also cell shape and migration, likely through a common regulatory mechanism on cytoskeletons.

Highlights

Cell migration is a highly orchestrated process throughout the embryonic development, and during adult wound healing, tissue repair and regeneration, immune responses, and tumor metastasis [1]
Mertk affects phagocytosis through the regulation of cytoskeletal rearrangement, which may be critical for cell attachment and spreading, we examined the effect of the loss of Mertk on the cell attachment on the culture dishes
Since cell migration depends on cytoskeletal rearrangement and rapid adhesion disassembly and assembly involving integrins and their downstream focal adhesion kinase (FAK), we examined the FAK phosphorylation on the Mertk-/- macrophages simulated with apoptotic cells for period times

Summary

Introduction

Cell migration is a highly orchestrated process throughout the embryonic development, and during adult wound healing, tissue repair and regeneration, immune responses, and tumor metastasis [1]. Formation of actin filements in lamellipodia and filopodia is regulated through a serial signaling cascade and the Rho family of small guanosine triphosphate (GTP)-binding proteins (GTPases), such as Rac, and Cdc, is critical for protrusion formation of lamellipodia and filopodia Both can activate downstream WAVE/WASP proteins that further stimulate the Arp2/3 complex to induce actin polymerization [1,5]. Another key signaling pathway that regulates cell migration, especially toward chemoatractant gradients, involves the PI3Ks and Pten phosphatase, which collaboratively regulate the local contents of phosphoinositides PI(3,4,5,)P3 and PI(3,4)P2. FAK is a key tyrosine kinase in linking integrins to downstream Rac-specific GEFs that in turn regulate actin polymerization or turnover [5]

Methods

Results

Conclusion