Podosomes and invadopodia, collectively known as invadosomes, are cell-matrix contacts in a variety of cell types, such as monocytic cells or cancer cells, that have to cross tissue barriers. Both structures share an actin-rich core, which distinguishes them from other matrix contacts, and are regulated by a multitude of signaling pathways including RhoGTPases, kinases, actin-associated proteins, and microtubule-dependent transport. Invadosomes recruit and secrete proteinases and are thus able to lyse extracellular matrix components. They are therefore considered to be potential key structures in proteolytic cell invasion in both physiological and pathological settings. This review provides an overview of the field, with special focus on current developments such as intracellular transport processes, ultrastructural analysis, the possible involvement of invadosomes in disease, and the tentative identification of invadosomes in 3D environments and in vivo.

This article is part of a Minifocus on invadopodia and podosomes. For further reading, please see related articles: `Matrix invasion by tumour cells: a focus on MT1-MMP trafficking to invadopodia' by Renaud Poincloux et al. (J. Cell Sci. 122 , [3015-3024][1]), \`Mechanisms for transcellular

Migration of macrophages is a key process for a variety of physiological functions, such as pathogen clearance or tissue homeostasis. However, it can also be part of pathological scenarios, as in the case of tumor-associated macrophages. This review presents an overview of the different migration modes macrophages can adopt, depending on the physical and chemical properties of specific environments, and the constraints they impose upon cells. We discuss the importance of these environmental and also of cellular parameters, as well as their relative impact on macrophage migration and on the formation of matrix-lytic podosomes in 2D and 3D. Moreover, we present an overview of routinely used and also newly developed assays for the study of macrophage migration in both 2D and 3D contexts, their respective advantages and limitations, and also their potential to reliably mimic in vivo situations.

Podosomes are actin-rich adhesion and invasion structures. Especially in macrophages, podosomes exist in two subpopulations, large precursors at the cell periphery and smaller podosomes (successors) in the cell interior. To date, the mechanisms that differentially regulate these subpopulations are largely unknown. Here, we show that the membrane-associated protein supervillin localizes preferentially to successor podosomes and becomes enriched at precursors immediately prior to their dissolution. Consistently, podosome numbers are inversely correlated with supervillin protein levels. Using deletion constructs, we find that the myosin II-regulatory N-terminus of supervillin (SV 1-174) is crucial for these effects. Phosphorylated myosin light chain (pMLC) localizes at supervillin-positive podosomes, and time-lapse analyses show that enrichment of GFP-supervillin at podosomes coincides with their coupling to contractile myosin IIA-positive cables. We also show that supervillin binds only to activated myosin IIA, and a dysregulated N-terminal construct (SV 1-830) enhances pMLC levels at podosomes. Thus, preferential recruitment of supervillin to podosome subpopulations may both require and induce actomyosin contractility. Using siRNA and pharmacological inhibition, we demonstrate that supervillin and myosin IIA cooperate to regulate podosome lifetime, podosomal matrix degradation and cell polarization. In sum, we show here that podosome subpopulations differ in their molecular composition and identify supervillin, in cooperation with myosin IIA, as a critical factor in the regulation of podosome turnover and function.