Figure 1. Elevated phospho-MARCKS abundance in invasive breast cancer.

A. Correlation of high phospho-MARCKS levels with distant metastasis of breast cancer. Left, representative images of immunohistochemical (IHC) staining in adjacent non-tumor areas and breast cancer specimens with low levels of phospho-MARCKS (score = +1 or +2) and high phospho-MARCKS abundance (score = +3). Right, percentage of patients with high and low phospho-MARCKS according to no distant metastasis (M0) vs distant metastasis (M1). P = 0.042, Fisher's exact test. B. Left, representative IHC images of breast primary tumors and lymph node metastatic tumors by using anti-phospho-MARCKS and anti-MARCKS antibodies. Right, percentage of patients with high and low phospho-MARCKS according to cancer types (noninvasive ductal carcinomas in situ, invasive breast carcinomas and lymph node metastatic tumors). p = 0.048, Fisher's exact test. (C-D) Higher phospho-MARCKS promotes breast cancer cell invasion and migration. MDA-MB-231 cells were transduced with control non-specific or MARCKS-specific shRNA-containing lentiviruses. Following knockdown of MARCKS, cells were re-expressed either wild type or mutant (S159/163A) V5-tagged MARCKS. C. Left, the levels of phospho-MARCKS abundance and MARCKS expression in these genetically modified cells were determined by Western blot. These cells were plated on Transwells with matrigel; 20 hours later, migrated cells were fixed, stained, and counted using light microscopy. A representative picture of each group is shown in the middle. Right, quantification of migrated cells to the lower chamber. Data expressed as mean ± SD (n = 4), *p < 0.05 as compared to cells receiving control shRNA. D. Confluent cultures of these cells were scratched and wound healing repair was monitored microscopically after the scratch. Left, representative phase contrast pictures. Right, numbers of cells migrated to the wound area were quantified at 16 hours post-scratching. (n = 4, *p < 0.05 versus control shRNA).