SUN-MKL1 Crosstalk Regulates Nuclear Deformation and Fast Motility of Breast Carcinoma Cells in Fibrillar ECM Microenvironment.

Ved P Sharma,David Entenberg,Robert Eddy,Maja H Oktay,Edison Leung,Joe Sanders,James Castracane,James Williams,John S Condeelis

doi:10.3390/cells10061549

Ved P Sharma, David Entenberg + Show 7 more

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https://doi.org/10.3390/cells10061549

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Abstract

Aligned collagen fibers provide topography for the rapid migration of single tumor cells (streaming migration) to invade the surrounding stroma, move within tumor nests towards blood vessels to intravasate and form distant metastases. Mechanisms of tumor cell motility have been studied extensively in the 2D context, but the mechanistic understanding of rapid single tumor cell motility in the in vivo context is still lacking. Here, we show that streaming tumor cells in vivo use collagen fibers with diameters below 3 µm. Employing 1D migration assays with matching in vivo fiber dimensions, we found a dependence of tumor cell motility on 1D substrate width, with cells moving the fastest and the most persistently on the narrowest 1D fibers (700 nm–2.5 µm). Interestingly, we also observed nuclear deformation in the absence of restricting extracellular matrix pores during high speed carcinoma cell migration in 1D, similar to the nuclear deformation observed in tumor cells in vivo. Further, we found that actomyosin machinery is aligned along the 1D axis and actomyosin contractility synchronously regulates cell motility and nuclear deformation. To further investigate the link between cell speed and nuclear deformation, we focused on the Linker of Nucleoskeleton and Cytoskeleton (LINC) complex proteins and SRF-MKL1 signaling, key regulators of mechanotransduction, actomyosin contractility and actin-based cell motility. Analysis of The Cancer Genome Atlas dataset showed a dramatic decrease in the LINC complex proteins SUN1 and SUN2 in primary tumor compared to the normal tissue. Disruption of LINC complex by SUN1 + 2 KD led to multi-lobular elongated nuclei, increased tumor cell motility and concomitant increase in F-actin, without affecting Lamin proteins. Mechanistically, we found that MKL1, an effector of changes in cellular G-actin to F-actin ratio, is required for increased 1D motility seen in SUN1 + 2 KD cells. Thus, we demonstrate a previously unrecognized crosstalk between SUN proteins and MKL1 transcription factor in modulating nuclear shape and carcinoma cell motility in an in vivo relevant 1D microenvironment.

Highlights

The tumor microenvironment (TME) plays an essential role in breast cancer invasion and metastasis [1,2,3,4,5]
In order to reconstitute in vitro tumor cell motility with high fidelity to the in vivo tumor cell motility phenotype, we first determined the in vivo tumor ECM architecture that is preferred by tumor cells during their migration
The MTLn3 and polyoma middle T antigen (PyMT) breast cancer mouse models are two of the best characterized mouse models of tumor cell invasion, motility and metastasis [30,49,50]. In these breast cancer models it is well established that tumor cells migrate on linear fibrillary ECM fibers [5,11,16,28,30,50]. Using these mammary tumor models, we imaged the fibrillar collagen I associated with tumor cell motility in vivo using second harmonic generation (SHG) intravital microscopy (Figure 1A,B, Movie S1)

Summary

Introduction

The tumor microenvironment (TME) plays an essential role in breast cancer invasion and metastasis [1,2,3,4,5]. TME consists of ECM, stromal cells (e.g., cancer associated fibroblasts, adipocytes) and immune cells (e.g., macrophages, neutrophils, etc.) [6,7,8], and drives breast tumor progression through mechanical and chemical cues [1,2,4,8]. The biomechanical properties of tumor ECM (mammographic breast density, ECM stiffness, crosslinking, topography and alignment) play important roles during breast tumor progression. Accumulation of collagen I is associated with increased risk of metastasis in breast cancer [12,13,14]. Striking changes in collagen I architecture (topography and alignment) are observed during breast tumor progression. Multiple studies have reported aligned collagen fibers oriented perpendicular to the tumor boundary in human breast tumor tissue sections [15,16]

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