Protein kinase G expression in the small intestine and functional importance for smooth muscle relaxation

Huber, Andrea; Trudrung, P.; Storr, Martin; Franck, H.; Schusdziarra, V.; Ruth, Peter; Allescher, Hans–Dieter

doi:10.1152/ajpgi.1998.275.4.g629

Cited by 24 publications

(25 citation statements)

References 45 publications

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“…In our study, the examination of the endothelium and nerve ®bers revealed no clear expression of cGK I, although cGK I expression was found in the nerve ®bers and endothelium of other organs. 20,21 Furthermore we have not found in our study any distinct differences between potent and impotent patients in regards to the distribution of sGC and cGMP dependent kinase I. We have to stress the limitations of our study regarding the small number of patients which did not allow a statistical analysis.…”

Section: Discussionmentioning

confidence: 63%

Soluble guanylate cyclase and cGMP-dependent protein kinase I expression in the human corpus cavernosum

et al. 2000

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Nitric oxide (NO) as a mediator in smooth muscle cells causes rapid and robust increases in cGMP levels. The cGMP-dependent protein kinase I has emerged as an important signal transduction mediator for smooth muscle relaxation. The purpose of this study was to examine the existence and distribution of two key enzymes of the NOacGMP pathway, the cGMP-dependent kinase I (cGK I) and the soluble guanylate cyclase (sGC) in human cavernosal tissue.The expression of the enzymes were examined in corpus cavernosum specimens of 23 patients. Eleven potent patients suffered from penile deviations and were treated via Nesbit's surgical method. Nine long-term impotent patients underwent implantation of¯exible hydraulic prothesis. Three potent patients underwent trans-sexual operations. Expression of the sGC and cGK I were examined immunohistochemically using speci®c antibodies.In all specimens of cavernosal tissue a distinct immunoreactivity was observed in different parts and structures. We found a high expression of sGC and cGK I in smooth muscle cells of vessels and in the ®bromuscular stroma. The endothelium of the cavernosal sinus, of the cavernosal arteries, and the cavernosal nerve ®bers showed an immunoreactivity against sGC. The distribution analysis of cGK I revealed a predominately vesicular localization in smooth muscle cells. The examination of the endothelium showed no clear immunoreactivity against cGK I. There was no distinct difference in immunoreactivity and cellular distribution between potent and impotent patients.cGMP-dependent kinase I is predominately expressed in cavernosal smooth muscle cells. sGC is expressed in smooth muscle cells, endothelium and nerve ®bers of the corpus cavernosum. In this study, there is no correlation between the distribution of soluble guanylate cyclase and cGMP dependent kinase I in potent or impotent patients.

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Section: Discussionmentioning

confidence: 63%

Soluble guanylate cyclase and cGMP-dependent protein kinase I expression in the human corpus cavernosum

et al. 2000

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“…This upstream component of the cGMP signaling pathway is targeted by sildenafil, a selective phosphodiesterase type 5 inhibitor that somewhat selectively regulates blood flow in the corpus cavernosum. Although some studies (67,68) have reported differences in the relative expression of the two cGKI isoforms that could influence the response to cGMP, no smooth muscle tissue has been identified that expresses one isoform exclusively. The lack of an antibody against chicken cGKI isoforms precluded a determination of cGKI isoform expression, but semi-quantitative RT-PCR indicated no significant change in cGKI␣ mRNA abundance during gizzard development (data not shown).…”

Section: Discussionmentioning

confidence: 99%

Role of Myosin Phosphatase Isoforms in cGMP-mediated Smooth Muscle Relaxation

Khatri¹,

Joyce²,

Brozovich³

et al. 2001

Journal of Biological Chemistry

115

178

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In vitro experiments showing the activation of the myosin phosphatase via heterophilic leucine zipper interactions between its targeting subunit (MYPT1) and cGMP-dependent protein kinase I suggested a pathway for smooth muscle relaxation (Surks, H. K., Mochizuki, N., Kasai, Y., Georgescu, S. P., Tang, K. M., Ito, M., Lincoln, T. M., and Mendelsohn, M. E. (1999) Science 286, 1583-1587). The relationship between MYPT1 isoform expression and smooth muscle responses to cGMP signaling in vivo has not been explored. MYPT1 isoforms that contain or lack a C-terminal leucine zipper are generated in birds and mammals by cassette-type alternative splicing of a 31-nucleotide exon. The avian and mammalian C-terminal isoforms are highly conserved and expressed in a tissue-specific fashion. In the mature chicken the tonic contracting aorta and phasic contracting gizzard exclusively express the leucine zipper positive and negative MYPT1 isoforms, respectively. Expression of the MYPT1 isoforms is also developmentally regulated in the gizzard, which switches from leucine zipper positive to negative isoforms around the time of hatching. This switch coincides with the development in the gizzard of a cGMP-resistant phenotype, i.e. inability to dephosphorylate myosin and relax in response to 8-bromo-cGMP after calcium activation. Furthermore, association of cGMP-dependent protein kinase I with MYPT1 is detected by immunoprecipitation only in the tissue that expresses the leucine zipper positive isoform of MYPT1. These results suggest that the regulated splicing of MYPT1 is an important determinant of smooth muscle phenotypic diversity and the variability in the response of smooth muscles to the calcium desensitizing effect of cGMP signaling.Smooth muscle contraction is initiated by the phosphorylation of the regulatory myosin light chain (MLC 20 ) 1 by the calcium/calmodulin-dependent activation of the myosin light chain kinase (MLCK) (1). Relaxation is effected by the dephosphorylation of MLC 20 by the smooth muscle myosin phosphatase (SMMP). Complexity is brought to this system by accessory proteins and signaling pathways that regulate the smooth muscle contractile state (reviewed in Refs. 2 and 3). The SMMP is a target of signals that are positive and negative modulators of smooth muscle tone. SMMP is a heterotrimeric protein composed of the 37-kDa catalytic subunit (PP1c␦), the 130/133-kDa myosin targeting subunit (MYPT1, also referred to as MBS), and the 21-kDa M21 subunit (4 -6). MYPT1 targets the catalytic subunit to MLC 20 (7,8) and in this way confers substrate specificity to the phosphatase, whereas the function of the M21 subunit is unknown. Activation of the Rho kinase signaling pathway leads to phosphorylation of the MYPT1 subunit, resulting in inhibition of myosin phosphatase activity and an increase in smooth muscle tone (9 -13). This signaling pathway is thought to determine the calcium-sensitizing effect of ␣-adrenergic stimulation, for example, in which greater force is produced at a given calcium concentration th...

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“…The inhibitory effect of NO on smooth muscle cells, either of the gut wall (5) or intestinal blood vessels (6), regulates gut motility and blood flow. Further, NO modulates neurosecretion from enteric nerve terminals (7).…”

mentioning

confidence: 99%

“…A major target of cGMP in smooth muscle is the cGMP-dependent protein kinase I (cGKI) 1 (12)(13)(14). The two existing isoforms of cGKI, cGKI␣ and cGKI␤, show a tissue-specific distribution (15), with cGKI␤ being the predominant isoform in the small intestine (5). The importance of the NO/cGMP signaling in the gut has been demonstrated by gene ablation.…”

mentioning

confidence: 99%

“…These proteins may be involved in relaxation and contractions of vascular smooth muscle by regulating cytoskeletal organization and cell shape. In contrast to vascular smooth muscle, little is known about potential targets of cGKI in the gastrointestinal tract, although the relaxing effect of cGKI on this type of smooth muscle is well described (5,27). In addition, the downstream effectors of cGKI in enteric neurons remains to be established.…”

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confidence: 99%

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Cysteine-rich Protein 2, a Novel Substrate for cGMP Kinase I in Enteric Neurons and Intestinal Smooth Muscle

Huber¹,

Neuhuber²,

Klugbauer³

et al. 2000

Journal of Biological Chemistry

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Nitric oxide/cGMP/cGMP kinase I (cGKI) signaling causes relaxation of intestinal smooth muscle. In the gastrointestinal tract substrates of cGKI have not been identified yet. In the present study a protein interacting with cGKI␤ has been isolated from a rat intestinal cDNA library using the yeast two-hybrid system. The protein was identified as cysteine-rich protein 2 (CRP2), recently cloned from rat brain (Okano, I., Yamamoto, T., Kaji, A., Kimura, T., Mizuno, K., and Nakamura, T. (1993) FEBS Lett. 333, 51-55). Recombinant CRP2 is specifically phosphorylated by cGKs but not by cAMP kinase in vitro. Co-transfection of CRP2 and cGKI␤ into COS cells confirmed the phosphorylation of CRP2 in vivo. Cyclic GMP kinase I phosphorylated CRP2 at Ser-104, because the mutation to Ala completely prevented the in vivo phosphorylation. Immunohistochemical analysis using confocal laser scan microscopy showed a co-localization of CRP2 and cGKI in the inner part of the circular muscle layer, in the muscularis mucosae, and in specific neurons of the myenteric and submucosal plexus. The co-localization together with the specific phosphorylation of CRP2 by cGKI in vitro and in vivo suggests that CRP2 is a novel substrate of cGKI in neurons and smooth muscle of the small intestine.Nitric oxide, a candidate of nonadrenergic, noncholinergic neurotransmission of the enteric nervous system (1, 2) has a variety of physiological functions in the gut (3, 4). The inhibitory effect of NO on smooth muscle cells, either of the gut wall (5) or intestinal blood vessels (6), regulates gut motility and blood flow. Further, NO modulates neurosecretion from enteric nerve terminals (7). A main signaling pathway of NO in the gut is the activation of soluble guanylyl cyclase (8 -10), which results in an increase of intracellular cGMP. Cyclic GMP analogues elicit NO-like effects on smooth muscle (11) and isolated nerve terminals (7) causing smooth muscle relaxation and release of vasoactive intestinal polypeptide, respectively. A major target of cGMP in smooth muscle is the cGMP-dependent protein kinase I (cGKI) 1 (12-14). The two existing isoforms of cGKI, cGKI␣ and cGKI␤, show a tissue-specific distribution (15), with cGKI␤ being the predominant isoform in the small intestine (5). (22) and on the stimulation of the large conductance K Ca channel (BK) (23,24). Furthermore, the relaxation of smooth muscle by NO/cGMP is also associated with an increase in the extent of phosphorylation of small proteins like heat shock related proteins that appear to be regulatory components of the actin based cytoskeleton or that seem to interact with intermediate filaments (25). A further substrate of cGKI involved in actin filament assembling and cell motility is the vasodilator-stimulated phosphoprotein associated with focal adhesions and areas of dynamic membrane activity (26). These proteins may be involved in relaxation and contractions of vascular smooth muscle by regulating cytoskeletal organization and cell shape. In contrast to vascular smooth muscle, little is...

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Protein kinase G expression in the small intestine and functional importance for smooth muscle relaxation

Cited by 24 publications

References 45 publications

Soluble guanylate cyclase and cGMP-dependent protein kinase I expression in the human corpus cavernosum

Soluble guanylate cyclase and cGMP-dependent protein kinase I expression in the human corpus cavernosum

Role of Myosin Phosphatase Isoforms in cGMP-mediated Smooth Muscle Relaxation

Cysteine-rich Protein 2, a Novel Substrate for cGMP Kinase I in Enteric Neurons and Intestinal Smooth Muscle

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