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24 pages, 6997 KiB

Open AccessArticle

Testosterone Enhances K_V Currents and Airway Smooth Muscle Relaxation Induced by ATP and UTP through P2Y₄ Receptors and Adenylyl Cyclase Pathway

by Abril Carbajal-García, Jorge Reyes-García, Verónica Díaz-Hernández, María F. Casas-Hernández, Francisco Javier Flores-Murrieta and Luis M. Montaño

Int. J. Mol. Sci. 2024, 25(9), 4652; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms25094652 - 24 Apr 2024

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Abstract

Numerous studies suggest the involvement of adenosine-5′-triphosphate (ATP) and similar nucleotides in the pathophysiology of asthma. Androgens, such as testosterone (TES), are proposed to alleviate asthma symptoms in young men. ATP and uridine-5′-triphosphate (UTP) relax the airway smooth muscle (ASM) via purinergic P2Y [...] Read more.

Numerous studies suggest the involvement of adenosine-5′-triphosphate (ATP) and similar nucleotides in the pathophysiology of asthma. Androgens, such as testosterone (TES), are proposed to alleviate asthma symptoms in young men. ATP and uridine-5′-triphosphate (UTP) relax the airway smooth muscle (ASM) via purinergic P2Y₂ and P2Y₄ receptors and K⁺ channel opening. We previously demonstrated that TES increased the expression of voltage-dependent K⁺ (K_V) channels in ASM. This study investigates how TES may potentiate ASM relaxation induced by ATP and UTP. Tracheal tissues treated with or without TES (control group) from young male guinea pigs were used. In organ baths, tracheas exposed to TES (40 nM for 48 h) showed enhanced ATP- and UTP-evoked relaxation. Tetraethylammonium, a K⁺ channel blocker, annulled this effect. Patch-clamp experiments in tracheal myocytes showed that TES also increased ATP- and UTP-induced K⁺ currents, and this effect was abolished with flutamide (an androgen receptor antagonist). K_V channels were involved in this phenomenon, which was demonstrated by inhibition with 4-aminopyridine. RB2 (an antagonist of almost all P2Y receptors except for P2Y₂), as well as N-ethylmaleimide and SQ 22,536 (inhibitors of G proteins and adenylyl cyclase, respectively), attenuated the enhancement of the K⁺ currents induced by TES. Immunofluorescence and immunohistochemistry studies revealed that TES did not modify the expression of P2Y₄ receptors or COX-1 and COX-2, while we have demonstrated that this androgen augmented the expression of K_V1.2 and K_V1.5 channels in ASM. Thus, TES leads to the upregulation of P2Y₄ signaling and K_V channels in guinea pig ASM, enhancing ATP and UTP relaxation responses, which likely limits the severity of bronchospasm in young males. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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23 pages, 4626 KiB

Open AccessArticle

A Conserved Intramolecular Ion-Pair Plays a Critical but Divergent Role in Regulation of Dimerization and Transport Function among the Monoamine Transporters

by Sixiang Chen, Xingyu Huang, Xintong Zhang, Chan Li and Yuan-Wei Zhang

Int. J. Mol. Sci. 2024, 25(7), 4032; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms25074032 - 04 Apr 2024

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Abstract

The monoamine transporters, including the serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET), are the therapeutic targets for the treatment of many neuropsychiatric disorders. Despite significant progress in characterizing the structures and transport mechanisms of these transporters, the regulation of their [...] Read more.

The monoamine transporters, including the serotonin transporter (SERT), dopamine transporter (DAT), and norepinephrine transporter (NET), are the therapeutic targets for the treatment of many neuropsychiatric disorders. Despite significant progress in characterizing the structures and transport mechanisms of these transporters, the regulation of their transport functions through dimerization or oligomerization remains to be understood. In the present study, we identified a conserved intramolecular ion-pair at the third extracellular loop (EL3) connecting TM5 and TM6 that plays a critical but divergent role in the modulation of dimerization and transport functions among the monoamine transporters. The disruption of the ion-pair interactions by mutations induced a significant spontaneous cross-linking of a cysteine mutant of SERT and an increase in cell surface expression but with an impaired specific transport activity. On the other hand, similar mutations of the corresponding ion-pair residues in both DAT and NET resulted in an opposite effect on their oxidation-induced dimerization, cell surface expression, and transport function. Reversible biotinylation experiments indicated that the ion-pair mutations slowed down the internalization of SERT but stimulated the internalization of DAT. In addition, cysteine accessibility measurements for monitoring SERT conformational changes indicated that substitution of the ion-pair residues resulted in profound effects on the rate constants for cysteine modification in both the extracellular and cytoplasmatic substrate permeation pathways. Furthermore, molecular dynamics simulations showed that the ion-pair mutations increased the interfacial interactions in a SERT dimer but decreased it in a DAT dimer. Taken together, we propose that the transport function is modulated by the equilibrium between monomers and dimers on the cell surface, which is regulated by a potential compensatory mechanism but with different molecular solutions among the monoamine transporters. The present study provided new insights into the structural elements regulating the transport function of the monoamine transporters through their dimerization. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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15 pages, 4538 KiB

Open AccessArticle

Deletion of Sarcolemmal Membrane-Associated Protein Isoform 3 (SLMAP3) in Cardiac Progenitors Delays Embryonic Growth of Myocardium without Affecting Hippo Pathway

by Taha Rehmani, Ana Paula Dias, Marsel Kamal, Maysoon Salih and Balwant S. Tuana

Int. J. Mol. Sci. 2024, 25(5), 2888; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms25052888 - 01 Mar 2024

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Abstract

The slmap gene is alternatively spliced to generate many isoforms that are abundant in developing myocardium. The largest protein isoform SLMAP3 is ubiquitously expressed and has been linked to cardiomyopathy, Brugada syndrome and Hippo signaling. To examine any role in cardiogenesis, mice homozygous [...] Read more.

The slmap gene is alternatively spliced to generate many isoforms that are abundant in developing myocardium. The largest protein isoform SLMAP3 is ubiquitously expressed and has been linked to cardiomyopathy, Brugada syndrome and Hippo signaling. To examine any role in cardiogenesis, mice homozygous for floxed slmap allele were crossed with Nkx2.5-cre mice to nullify its expression in cardiac progenitors. Targeted deletion of the slmap gene resulted in the specific knockout (KO) of the SLMAP3 (~91 KDa) isoform without any changes in the expression of the SLMAP2 (~43 kDa) or the SLMAP1 (~35 kDa) isoforms which continued to accumulate to similar levels as seen in Wt embryonic hearts. The loss of SLMAP3 from cardiac progenitors resulted in decreased size of the developing embryonic hearts evident at E9.5 to E16.5 with four small chambers and significantly thinner left ventricles. The proliferative capacity assessed with the phosphorylation of histone 3 or with Ki67 in E12.5 hearts was not significantly altered due to SLMAP3 deficiency. The size of embryonic cardiomyocytes, marked with anti-Troponin C, revealed significantly smaller cells, but their hypertrophic response (AKT1 and MTOR1) was not significantly affected by the specific loss of SLMAP3 protein. Further, no changes in phosphorylation of MST1/2 or YAP were detected in SLMAP3-KO embryonic myocardium, ruling out any impact on Hippo signaling. Rat embryonic cardiomyocytes express the three SLMAP isoforms and their knockdown (KD) with sh-RNA, resulted in decreased proliferation and enhanced senescence but without any impact on Hippo signaling. Collectively, these data show that SLMAP is critical for normal cardiac development with potential for the various isoforms to serve compensatory roles. Our data imply novel mechanisms for SLMAP action in cardiac growth independent of Hippo signaling. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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11 pages, 3384 KiB

Open AccessArticle

Effect of THz Waves of Different Orientations on K⁺ Permeation Efficiency in the KcsA Channel

by Yize Wang, Hongguang Wang, Wen Ding, Xiaofei Zhao, Yongdong Li and Chunliang Liu

Int. J. Mol. Sci. 2024, 25(1), 429; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms25010429 - 28 Dec 2023

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Abstract

Potassium (K) channels show the highest variability and most frequent alterations in expression in many tumor types, and modulation of K⁺ channels may represent a new window for cancer therapy. In previous work, we found that a terahertz (THz) field incident along [...] Read more.

Potassium (K) channels show the highest variability and most frequent alterations in expression in many tumor types, and modulation of K⁺ channels may represent a new window for cancer therapy. In previous work, we found that a terahertz (THz) field incident along the z-axis with a frequency of 51.87 THz increased the ion flux through K⁺ channels. In practice, it is difficult to ensure that the incident electromagnetic (EM) wave is strictly parallel to the direction of channel ion flow. In this paper, we found by changing the direction of the applied electric field that the EM wave of a specific frequency has the largest ion flux when the incident direction is along the ion flow, and the smallest ion flux when the incident direction is perpendicular to the ion flow, and that overall the EM wave of this frequency enhances the ion flow of the K⁺ channel. Changes in the direction of the applied field at a specific frequency affect the stability of the φ dihedral angle of the GLY77 residue and alter the ion permeation mechanism in the selectivity filter (SF) region, thus affecting the ion flux. Therefore, this frequency can be used to modulate K⁺ fluxes by THz waves to cause rapid apoptosis in potassium-overloaded tumor cells. This approach consequently represents an important tool for the treatment of cancer and is expected to be applied in practical therapy. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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13 pages, 5825 KiB

Open AccessArticle

Host-Cell-Dependent Roles of E-Cadherin in Serratia Invasion

by Olga Tsaplina, Ekaterina Lomert and Yuliya Berson

Int. J. Mol. Sci. 2023, 24(23), 17075; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms242317075 - 03 Dec 2023

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Abstract

Bacteria use cell surface proteins to mediate host–pathogen interactions. Proteins responsible for cell adhesion, including E-cadherin, serve as receptors for entry into the host cell. We have previously shown that an increase in eukaryotic cell sensitivity to Serratia grimesii correlates with an increase [...] Read more.

Bacteria use cell surface proteins to mediate host–pathogen interactions. Proteins responsible for cell adhesion, including E-cadherin, serve as receptors for entry into the host cell. We have previously shown that an increase in eukaryotic cell sensitivity to Serratia grimesii correlates with an increase in E-cadherin expression. On the other hand, Serratia proteamaculans invasion involves the EGFR, which can interact with E-cadherin on the surface of host cells. Therefore, we investigated the role of E-cadherin in Serratia invasion into M-HeLa and Caco-2 cells. Bacterial infection increased E-cadherin expression in both cell lines. Moreover, E-cadherin was detected in the Caco-2 cells in a full-length form and in the M-HeLa cells in only a truncated form in response to incubation with bacteria. Transfection with siRNA targeting E-cadherin inhibited S. proteamaculans invasion only into the Caco-2 cells. Thus, only full-length E-cadherin is involved in S. proteamaculans invasion. On the other hand, transfection with siRNA targeting E-cadherin inhibited S. grimesii invasion into both cell lines. Thus, not only may full-length E-cadherin but also truncated E-cadherin be involved in S. grimesii invasion. Truncated E-cadherin can be formed as a result of cleavage by bacterial proteases or the Ca²⁺-activated cellular protease ADAM10. The rate of Ca²⁺ accumulation in the host cells depends on the number of bacteria per cell upon infection. During incubation, Ca²⁺ accumulates only when more than 500 S. grimesii bacteria are infected per eukaryotic cell, and only under these conditions does the ADAM10 inhibitor reduce the sensitivity of the cells to bacteria. An EGFR inhibitor has the same quantitative effect on S. grimesii invasion. Apparently, as a result of infection with S. grimesii, Ca²⁺ accumulates in the host cells and may activate the ADAM10 sheddase, which can promote invasion by cleaving E-cadherin and, as a result, triggering EGFR signaling. Thus, the invasion of S. proteamaculans can only be promoted by full-length E-cadherin, and S. grimesii invasion can be promoted by both full-length and truncated E-cadherin. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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17 pages, 4147 KiB

Open AccessArticle

Ahf-Caltide, a Novel Polypeptide Derived from Calpastatin, Protects against Oxidative Stress Injury by Stabilizing the Expression of Ca_V1.2 Calcium Channel

by Yingchun Xue, Shi Zhou, Ling Yan, Yuelin Li, Xingrong Xu, Xianghui Wang, Etsuko Minobe, Masaki Kameyama, Liying Hao and Huiyuan Hu

Int. J. Mol. Sci. 2023, 24(21), 15729; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms242115729 - 29 Oct 2023

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Abstract

Reperfusion after ischemia would cause massive myocardial injury, which leads to oxidative stress (OS). Calcium homeostasis imbalance plays an essential role in myocardial OS injury. Ca_V1.2 calcium channel mediates calcium influx into cardiomyocytes, and its activity is modulated by a region [...] Read more.

Reperfusion after ischemia would cause massive myocardial injury, which leads to oxidative stress (OS). Calcium homeostasis imbalance plays an essential role in myocardial OS injury. Ca_V1.2 calcium channel mediates calcium influx into cardiomyocytes, and its activity is modulated by a region of calpastatin (CAST) domain L, CS_L54-64. In this study, the effect of Ahf-caltide, derived from CS_L54-64, on myocardial OS injury was investigated. Ahf-caltide decreased the levels of LDH, MDA and ROS and increased heart rate, coronary flow, cell survival and SOD activity during OS. In addition, Ahf-caltide permeated into H9c2 cells and increased Ca_V1.2, Ca_Vβ2 and CAST levels by inhibiting protein degradation. At different Ca²⁺ concentrations (25 nM, 10 μM, 1 mM), the binding of CS_L to the IQ motif in the C terminus of the Ca_V1.2 channel was increased in a H₂O₂ concentration-dependent manner. CS_L54-64 was predicted to be responsible for the binding of CS_L to Ca_V1.2. In conclusion, Ahf-caltide exerted a cardioprotective effect on myocardial OS injury by stabilizing Ca_V1.2 protein expression. Our study, for the first time, proposed that restoring calcium homeostasis by targeting the Ca_V1.2 calcium channel and its regulating factor CAST could be a novel treatment for myocardial OS injury. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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Review

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15 pages, 1417 KiB

Open AccessReview

TRP Ion Channels in Immune Cells and Their Implications for Inflammation

by Qiyue Yan, Chuanzhou Gao, Mei Li, Rui Lan, Shaohan Wei, Runsong Fan and Wei Cheng

Int. J. Mol. Sci. 2024, 25(5), 2719; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms25052719 - 27 Feb 2024

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Abstract

The transient receptor potential (TRP) ion channels act as cellular sensors and mediate a plethora of physiological processes, including somatosensation, proliferation, apoptosis, and metabolism. Under specific conditions, certain TRP channels are involved in inflammation and immune responses. Thus, focusing on the role of [...] Read more.

The transient receptor potential (TRP) ion channels act as cellular sensors and mediate a plethora of physiological processes, including somatosensation, proliferation, apoptosis, and metabolism. Under specific conditions, certain TRP channels are involved in inflammation and immune responses. Thus, focusing on the role of TRPs in immune system cells may contribute to resolving inflammation. In this review, we discuss the distribution of five subfamilies of mammalian TRP ion channels in immune system cells and how these ion channels function in inflammatory mechanisms. This review provides an overview of the current understanding of TRP ion channels in mediating inflammation and may offer potential avenues for therapeutic intervention. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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24 pages, 5317 KiB

Open AccessReview

Role of STIM1 in the Regulation of Cardiac Energy Substrate Preference

by Panpan Liu, Zhuli Yang, Youjun Wang and Aomin Sun

Int. J. Mol. Sci. 2023, 24(17), 13188; https://0-doi-org.brum.beds.ac.uk/10.3390/ijms241713188 - 25 Aug 2023

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Abstract

The heart requires a variety of energy substrates to maintain proper contractile function. Glucose and long-chain fatty acids (FA) are the major cardiac metabolic substrates under physiological conditions. Upon stress, a shift of cardiac substrate preference toward either glucose or FA is associated [...] Read more.

The heart requires a variety of energy substrates to maintain proper contractile function. Glucose and long-chain fatty acids (FA) are the major cardiac metabolic substrates under physiological conditions. Upon stress, a shift of cardiac substrate preference toward either glucose or FA is associated with cardiac diseases. For example, in pressure-overloaded hypertrophic hearts, there is a long-lasting substrate shift toward glucose, while in hearts with diabetic cardiomyopathy, the fuel is switched toward FA. Stromal interaction molecule 1 (STIM1), a well-established calcium (Ca²⁺) sensor of endoplasmic reticulum (ER) Ca²⁺ store, is increasingly recognized as a critical player in mediating both cardiac hypertrophy and diabetic cardiomyopathy. However, the cause–effect relationship between STIM1 and glucose/FA metabolism and the possible mechanisms by which STIM1 is involved in these cardiac metabolic diseases are poorly understood. In this review, we first discussed STIM1-dependent signaling in cardiomyocytes and metabolic changes in cardiac hypertrophy and diabetic cardiomyopathy. Second, we provided examples of the involvement of STIM1 in energy metabolism to discuss the emerging role of STIM1 in the regulation of energy substrate preference in metabolic cardiac diseases and speculated the corresponding underlying molecular mechanisms of the crosstalk between STIM1 and cardiac energy substrate preference. Finally, we briefly discussed and presented future perspectives on the possibility of targeting STIM1 to rescue cardiac metabolic diseases. Taken together, STIM1 emerges as a key player in regulating cardiac energy substrate preference, and revealing the underlying molecular mechanisms by which STIM1 mediates cardiac energy metabolism could be helpful to find novel targets to prevent or treat cardiac metabolic diseases. Full article

(This article belongs to the Special Issue Ion Movements and Membrane Proteins)

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