Two‐dimensional (2D) in vitro culture systems have for a number of years provided a controlled and versatile environment for the study of cell adhesion and migration, two interrelated cell functions critical to cancer metastasis. However, the organization and functions of focal adhesion proteins in cells embedded in physiologically more relevant 3D matrices is qualitatively and functionally different from their organization and functions on conventional 2D planar substrates. In a 3D, crosslinked, fibrillar collagen matrix, cell migration and protrusion activity are still regulated by focal adhesion proteins, such as pl30Cas, FAK, Zyxin, Vinculin, Talin, and VASP, but differently from the 2D case. This talk will describe the implications of the dependence of focal adhesion rotein‐based cellular functions on microenvironmental dimensionality in cancer. We will discuss the implications of this work in cancer metastasis.