scholarly journals ZIPK is critical for the motility and contractility of VSMCs through the regulation of nonmuscle myosin II isoforms

2014 ◽  
Vol 306 (9) ◽  
pp. H1275-H1286 ◽  
Author(s):  
Satoshi Komatsu ◽  
Mitsuo Ikebe
Keyword(s):  
Protein Kinase ◽  
Collagen Gel ◽  
Collagen Matrix ◽  
Short Hairpin Rna ◽  
Nonmuscle Myosin ◽  
Hairpin Rna ◽  
Interacting Protein ◽  
Collagen Gel Contraction ◽  
Zipper Interacting Protein Kinase ◽  
Short Hairpin

Migration of medial vascular smooth muscle cells (VSMCs) into the intimal layer contributes to pathological remodeling of the blood vessel in arterial hypertension and atherosclerosis. It is well established that reorganization of cytoskeletal networks is an essential component of cellular motile events. However, there is currently a lack of insight into the cellular characteristics of VSMC migration under three-dimensional environments. Here, we investigated the mechanisms of VSMC migration and remodeling using two different collagen matrix assays as in vitro models: migration of VSMCs within a collagen matrix for VSMC invasion and contraction of a collagen gel by VSMCs for VSMC remodeling and contraction. We found that nonmuscle myosin IIA (NMIIA) and nonmuscle myosin IIB (NMIIB) differentially contribute to the migratory capacity of VSMCs via NMII isoform-dependent cytoskeletal reorganization. Depletion of NMIIA by short hairpin RNA revealed a unique interplay between actomyosin and microtubules during VSMC migration. On the other hand, NMIIB was required for the structural maintenance of migrating VSMC. Interestingly, there was a significant difference between NMIIA and NMIIB knockdown in the VSMC migration but not in the VSMC-mediated collagen gel contraction. Furthermore, depletion of zipper-interacting protein kinase by short hairpin RNA resulted in an impairment of VSMC migration and a substantial decrease of VSMC-mediated collagen gel contraction. These results suggest that NMIIA and NMIIB uniquely control VSMC migration and may contribute to vascular remodeling, which are both regulated by zipper-interacting protein kinase.

scholarly journals Regulation of membrane KCNQ1/KCNE1 channel density by sphingomyelin synthase 1

2016 ◽  
Vol 311 (1) ◽  
pp. C15-C23 ◽  
Author(s):  
Meikui Wu ◽  
Makoto Takemoto ◽  
Makoto Taniguchi ◽  
Toru Takumi ◽  
Toshiro Okazaki ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Current Density ◽  
Short Hairpin Rna ◽  
Protein Kinase D ◽  
Dependent Manner ◽  
Hairpin Rna ◽  
Channel Density ◽  
Sphingomyelin Synthase ◽  
Short Hairpin ◽  
Channel Properties

Sphingomyelin synthase (SMS) catalyzes the conversion of phosphatidylcholine and ceramide to sphingomyelin and diacylglycerol. We previously showed that SMS1 deficiency leads to a reduction in expression of the K+ channel KCNQ1 in the inner ear (Lu MH, Takemoto M, Watanabe K, Luo H, Nishimura M, Yano M, Tomimoto H, Okazaki T, Oike Y, and Song WJ. J Physiol 590: 4029–4044, 2012), causing hearing loss. However, it remains unknown whether this change in expression is attributable to a cellular process or a systemic effect in the knockout animal. Here, we examined whether manipulation of SMS1 activity affects KCNQ1/KCNE1 currents in individual cells. To this end, we expressed the KCNQ1/KCNE1 channel in human embryonic kidney 293T cells and evaluated the effect of SMS1 manipulations on the channel using whole cell recording. Application of tricyclodecan-9-yl-xanthogenate, a nonspecific inhibitor of SMSs, significantly reduced current density and altered channel voltage dependence. Knockdown of SMS1 by a short hairpin RNA, however, reduced current density alone. Consistent with this, overexpression of SMS1 increased the current density without changing channel properties. Furthermore, application of protein kinase D inhibitors also suppressed current density without changing channel properties; this effect was nonadditive with that of SMS1 short hairpin RNA. These results suggest that SMS1 positively regulates KCNQ1/KCNE1 channel density in a protein kinase D-dependent manner.

Folate-linked lipoplexes for short hairpin RNA targeting claudin-3 delivery in ovarian cancer xenografts

2013 ◽  
Vol 172 (3) ◽  
pp. 679-689 ◽  
Author(s):  
Zhi-Yao He ◽  
Xia-Wei Wei ◽  
Min Luo ◽  
Shun-Tao Luo ◽  
Yang Yang ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Short Hairpin Rna ◽  
Hairpin Rna ◽  
Rna Targeting ◽  
Short Hairpin

Ca2+-independent contraction of longitudinal ileal smooth muscle is potentiated by a zipper-interacting protein kinase pseudosubstrate peptide

2009 ◽  
Vol 297 (2) ◽  
pp. G361-G370 ◽  
Author(s):  
Eikichi Ihara ◽  
Lori Moffat ◽  
Meredith A. Borman ◽  
Jennifer E. Amon ◽  
Michael P. Walsh ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Conformational Changes ◽  
Kinase Inhibitor ◽  
Smooth Muscles ◽  
Dominant Negative ◽  
Myosin Phosphatase ◽  
Interacting Protein ◽  
Zipper Interacting Protein Kinase

As a regulator of smooth muscle contraction, zipper-interacting protein kinase (ZIPK) can directly phosphorylate the myosin regulatory light chains (LC20) and produce contractile force. Synthetic peptides (SM-1 and AV25) derived from the autoinhibitory region of smooth muscle myosin light chain kinase can inhibit ZIPK activity in vitro. Paradoxically, treatment of Triton-skinned ileal smooth muscle strips with AV25, but not SM-1, potentiated Ca2+-independent, microcystin- and ZIPK-induced contractions. The AV25-induced potentiation was limited to ileal and colonic smooth muscles and was not observed in rat caudal artery. Thus the potentiation of Ca2+-independent contractions by AV25 appeared to be mediated by a mechanism unique to intestinal smooth muscle. AV25 treatment elicited increased phosphorylation of LC20 (both Ser-19 and Thr-18) and myosin phosphatase-targeting subunit (MYPT1, inhibitory Thr-697 site), suggesting involvement of a Ca2+-independent LC20 kinase with coincident inhibition of myosin phosphatase. The phosphorylation of the inhibitor of myosin phosphatase, CPI-17, was not affected. The AV25-induced potentiation was abolished by pretreatment with staurosporine, a broad-specificity kinase inhibitor, but specific inhibitors of Rho-associated kinase, PKC, and MAPK pathways had no effect. When a dominant-negative ZIPK [kinase-dead ZIPK(1–320)-D161A] was added to skinned ileal smooth muscle, the potentiation of microcystin-induced contraction by AV25 was blocked. Furthermore, pretreatment of skinned ileal muscle with SM-1 abolished AV25-induced potentiation. We conclude, therefore, that, even though AV25 is an in vitro inhibitor of ZIPK, activation of the ZIPK pathway occurs following application of AV25 to permeabilized ileal smooth muscle. Finally, we propose a mechanism whereby conformational changes in the pseudosubstrate region of ZIPK permit augmentation of ZIPK activity toward LC20 and MYPT1 in situ. AV25 or molecules based on its structure could be used in therapeutic situations to induce contractility in diseases of the gastrointestinal tract associated with hypomotility.

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