Neuronal calcium sensor 1 (NCS1), a calcium-binding protein, and transient receptor potential V4 (TRPV4), a plasma membrane calcium channel, are fundamental in the regulation of calcium homeostasis. The interactions of these proteins and their regulation by paclitaxel (PTX) were investigated using biochemical, pharmacological, and electrophysiological approaches in both a breast cancer epithelial cell model and a neuronal model. TRPV4 and NCS1 reciprocally immunoprecipitated each other, suggesting that they comprise a signaling complex. The functional consequence of this physical association was that TRPV4 currents increased with increased NCS1 expression. Calcium fluxes through TRPV4 correlated with the magnitude of TRPV4 currents and these calcium fluxes depended on NCS1 expression levels. Exposure to PTX amplified the acute effects of TRPV4 expression, currents, and calcium fluxes, but decreased the expression of NCS1. These findings augment the understanding of the properties of TRPV4, the role of NCS1 in the regulation of TRPV4, and the cellular mechanisms of PTX-induced neuropathy. TRPV4 and NCS1 physically and functionally interact. Increased expression of NCS1 enhances TRPV4 dependent currents, which are further amplified by treatment with the chemotherapeutic drug, paclitaxel, an effect associated with adverse effects of chemotherapy, including neuropathy.