The β-subunits of voltage-gated Ca 2+ (Ca V ) channels regulate the functional expression and several biophysical properties of highvoltage-activated Ca V channels. We find that Ca V β-subunits also determine channel regulation by the membrane phospholipid phosphatidylinositol 4,5-bisphosphate (PIP 2 ). When Ca V 1.3, -2.1, or -2.2 channels are cotransfected with the β3-subunit, a cytosolic protein, they can be inhibited by activating a voltage-sensitive lipid phosphatase to deplete PIP 2 . When these channels are coexpressed with a β2a-subunit, a palmitoylated peripheral membrane protein, the inhibition is much smaller. PIP 2 sensitivity could be increased by disabling the two palmitoylation sites in the β2a-subunit. To further test effects of membrane targeting of Ca V β-subunits on PIP 2 regulation, the N terminus of Lyn was ligated onto the cytosolic β3-subunit to confer lipidation. This chimera, like the Ca V β2a-subunit, displayed plasma membrane localization, slowed the inactivation of Ca V 2.2 channels, and increased the current density. In addition, the Lyn-β3 subunit significantly decreased Ca V channel inhibition by PIP 2 depletion. Evidently lipidation and membrane anchoring of Ca V β-subunits compete with the PIP 2 regulation of high-voltage-activated Ca V channels. Compared with expression with Ca V β3-subunits alone, inhibition of Ca V 2.2 channels by PIP 2 depletion could be significantly attenuated when β2a was coexpressed with β3. Our data suggest that the Ca V currents in neurons would be regulated by membrane PIP 2 to a degree that depends on their endogenous β-subunit combinations.M1 muscarinic receptor | voltage-sensing phosphatase V oltage-gated Ca 2+ (Ca V ) channels define excitable cells. These channels contribute to electrical excitability and Ca 2+ -dependent processes, such as neurotransmitter release, motility, and gene expression (1). An intriguing aspect of Ca V channels is their slow, voltage-independent modulation by G q protein-coupled receptors (G q PCRs) (2-5). The several proposed mechanisms for this slow modulation of Ca V channels include depletion of phosphatidylinositol 4,5-bisphosphate (PIP 2 ) via activation of phospholipase C (3-10), phosphorylation by protein kinases (11, 12), generation of arachidonic acid by phospholipase A 2 (13,14), and other unspecified pathways involving Gα q or 16). In the nervous system, the dissection of G q PCR modulation of Ca 2+ channels is made difficult by the generation of several second messengers and by the existence of multiple subtypes of Ca V channels.Ca V channels are a complex of three protein subunits. The α1-subunit forms the voltage-sensitive, Ca 2+ -selective pore and is the target of selective antagonists. The other auxiliary subunits shape the physiological properties of the α1-subunit. Here we emphasize β-subunits. These subunits associate tightly with the cytoplasmic domain of the α1-subunit, imparting increased peak currents, altered rates of activation and inactivation, and increased density of functional channels (17-...

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Tumor-associated macrophages (TAMs) are involved in tumor progression by promoting epithelial-mesenchymal transition (EMT), tumor cell invasion, migration and angiogenesis. However, in breast cancer, the clinical relevance of the TAM infiltration according to distinct histologic locations (intratumoral vs. stromal) and hormone receptor status is unclear. We investigated the significance of the levels of TAM infiltration in distinct histologic locations in invasive breast cancer. We also examined the relationship of the TAM levels with the clinicopathologic features of tumors, expression of EMT markers, and clinical outcomes. Finally, we analyzed the prognostic value of TAM levels according to hormone receptor status. High levels of infiltration of intratumoral, stromal and total TAMs were associated with high histologic grade, p53 overexpression, high Ki-67 proliferation index and negative hormone receptor status. Infiltration of TAMs was also correlated with overexpression of vimentin, smooth muscle actin and alteration of β-catenin. Overall, a high level of infiltration of intratumoral TAMs was associated with poor disease-free survival, and was found to be an independent prognostic factor. In subgroup analyses by hormone receptor status, a high level of infiltration of intratumoral TAM was an independent prognostic factor in the hormone receptor-positive subgroup, but not in the hormone-receptor negative subgroup. Our findings suggest that intratumoral TAMs play an important role in tumor progression in breast cancer, especially in the hormone receptor-positive group, and the level of TAM infiltration may be used as a prognostic factor and even a therapeutic target in breast cancer.