Hydrogen Sulfide

Highlights

• Under ER stress the CSE enzyme starts metabolising Cysteine instead of Cystathionine to produce H₂S. (R1)

• CBS deficiency also leads to chronic increase in H₂S production (R1)

• Excess of Hydrogen sulfide is implicated in lowered PPI which can be seen in Schizophrenia and Tourette Syndrome. (R2)

• Increased production of H₂S leads to prolonged sulfhydration of Akt protein, which deactivates it, leading to diminished ability of AKT to deactivate GSK3b by phosphorylation (R5)

• Thioredoxins are involved in H₂S production by MPST enzyme (R3)

• Significantly lower plasma H₂S was found in patients with Depression (R4)

Role of H2S in health

Hydrogen sulfide (H₂S) is a member of the gasotransmitter family that is associated with the maintenance of neuronal plasticity, excitability, and homeostatic functions.

It is mainly produced by the enzyme cystathionine-β-synthase (CBS) in the brain and the enzyme cystathionine-γ-lyase (CSE) in the peripheral tissues.

Abe and Kimura first demonstrated the influences of H₂S on synaptic plasticity. They showed that physiological concentrations of H₂S facilitated the induction of hippocampal LTP by increasing the activity of NMDARs. Inhibition of H₂S generation would lead to a reduction in NMDAR-mediated synaptic response and cause an impairment of LTP in the amygdala. (R4)

Prepulse Inhibition

Prepulse inhibition (PPI) is a neurological phenomenon in which a weaker prestimulus (prepulse) inhibits the reaction of an organism to a subsequent strong reflex-eliciting stimulus (pulse), often using the startle reflex. The stimuli are usually acoustic, but tactile stimuli (e.g. via air puffs onto the skin) and light stimuli are also used. When prepulse inhibition is high, the corresponding one-time startle response is reduced. Wikipedia

Disruptions of PPI are studied in humans and many other species. The most studied are deficits of PPI in schizophrenia, although this disease is not the only one to be associated with such deficits. They have been noted in panic disorder (Ludewig, et al., 2005), schizotypal personality disorder, obsessive-compulsive disorder (Swerdlow et al., 1993), Huntington’s disease, nocturnal enuresis and attention deficit disorder (Ornitz et al. 1992), and Tourette’s syndrome (R6; R7).

According to one study, people who have temporal lobe epilepsy with psychosis also show decreases in PPI, unlike those who have TLE without psychosis.[23] Therefore, PPI deficits are not typical to specific disease, but rather tell of disruptions in a specific brain circuit.

Chronic overproduction of H2S and increased oxidative stress

When CBS enzyme is deficient and can’t produce enough cystathionine, the downstream CSE enzyme switches to production of H₂S from cysteine. (R1)

Increased production of H₂S is supposed to amplify Nrf2 signalling by inhibiting Keap1 by sulfhydration.

At the same time increased sulfhydration of Akt protein deactivates it and impairs normal cellular response to upstream stimuli, such as insulin.

This leads to insulin resistance and impairs timely deactivation of GSK3b.

GSK3b is active under normal condition and deactivated by phosphorylation by Akt. However when Akt is sulfhydrated it can’t bind to GSK3b and GSK3b remains active.

Active GSK3b prevents Nrf2 signalling cascade by phosphorylating Nrf2 protein.

Therefore, chronically increased production of H₂S due to insufficient work of CBS leads to reduced Nrf2 and chronically increased oxidative stress.

CBS activity directly depends on Pyridoxal 5'-phosphate, heme b and L-Serine availability. Hence B6 or Iron deficiency are the most common causes of poor CBS function, along with high psychological stress, because cortisol reduces CBS transcription (R9).

Inhibits glycolysis and affects immune response

Our data suggest that excessive H2S produced by the infected WT mice reduce HIF-1α levels, thereby suppressing glycolysis and production of IL-1β, IL-6, and IL-12, and increasing bacterial burden.

Clinical relevance was demonstrated by the spatial distribution of H2S-producing enzymes in human necrotic, nonnecrotic, and cavitary pulmonary tuberculosis (TB) lesions.

In summary, CSE exacerbates TB pathogenesis by altering immunometabolism in mice and inhibiting CSE or modulating glycolysis are potential targets for host-directed TB control. (R20)

Protective role

H2S protects neurons by increasing the intracellular levels of GSH through enhancing the activity of xCT as well as glutamate cysteine ligase, a rate‐limiting enzyme for GSH production.

H2S also facilitates the cysteine transport possibly through the excitatory amino acid transporter EAAT3 (SLC1A1), which was initially identified as transporting aspartate and glutamate, but not through the alanine/serine/cysteine (ASC) transporter.

On the other hand, H2Sn protects cells by activating a Kelch ECH‐associating protein 1 (Keap1)/nuclear factor erythroid 2‐related factor 2 (Nrf2) pathway through the sulfuration of Keap1, leading to the activation of antioxidant genes including those for GSH production. (R13)

MPST interacts with NFS1 and MOCS3

Among these sulfurtransferases is the human mercaptopyruvate sulfurtransferase (MPST) also designated as tRNA thiouridine modification protein (TUM1).

The role of the human TUM1 protein has been suggested in a wide range of physiological processes in the cell among which are but not limited to involvement in Molybdenum cofactor (Moco) biosynthesis, cytosolic tRNA thiolation and generation of H2S as signaling molecule both in mitochondria and the cytosol.

Previous interaction studies showed that TUM1/MPST interacts with the L-cysteine desulfurase NFS1 and the Molybdenum cofactor biosynthesis protein 3 (MOCS3).

Here, we show that TUM1 is involved in the sulfur transfer for Molybdenum cofactor synthesis and tRNA thiomodification by spectrophotometric measurement of the activity of sulfite oxidase and liquid chromatography quantification of the level of sulfur-modified tRNA. Further, we show that TUM1 has a role in hydrogen sulfide production and cellular bioenergetics. (R12)

Detoxification of H₂S

Excessive level of H₂S reverses Electron Transport Chain at Complex II and inhibits Complex IV

We have discovered that at high H₂S concentrations, which are known to inhibit complex IV, a new redox cycle is established between SQOR and complex II, operating in reverse. Under these conditions, the purine nucleotide cycle and the malate aspartate shuttle furnish fumarate, which supports complex II reversal and leads to succinate accumulation." (R10)

Highlights

• H2S inhibits Complex IV
• Complex II works in reverse
• Malate-Aspartate shuttle furnish fumarate
• Succinate accumulates

CQ10 pool is a limiting factor for H₂S clearance

These results indicate that the CoQ pool limits sulfide clearance and, in the absence of competition from complex I, cells clear sulfide more efficiently. The data also reveal that complex II has the opposite effect, i.e., it is advantageous for sulfide clearance, consistent with our model that complex II reversal supports H₂S oxidation by catalyzing CoQH2 oxidation. (R10)

It seems that inhibition of Complex I might be beneficial for H₂S clearance.

Hcy and H₂S regulate CBS and CSE

Our results revealed that Na₂S treatment suppresses SP1 in a dose dependent manner, suggesting that H₂S may downregulate CTH by inhibiting SP1.

Moreover, our luciferase reporter assay on CTH 3′UTR demonstrated that miR-133a targets CTH. Since H₂S upregulates miR-133a, it could be an indirect mechanism for H₂S-mediated regulation of CTH.

In conclusion, H₂S and Hcy regulate CBS and CTH in a dose dependent manner.

CBS negatively regulates CTH.

H₂S upregulates CBS but downregulates CTH, possibly by suppressing SP1 and inducing miR-133a in cardiomyocytes. (R11)

CBS and CSE can translocate to mitochondria

CSE and CBS, are predominantly cytosolic, but they do translocate to the mitochondria as well.

In vascular smooth muscle cells, calcium influx triggers mitochondrial translocation of CSE, a process dependent on translocase of the outer membrane 20 (Tom20) to generate H₂S in the mitochondria.

The existence of CSE in the mitochondrial compartment was also suggested by earlier studies which report an increase in cystathionine content in rat liver mitochondria treated with propargylglycine, an inhibitor of CSE.

CBS, too is associated with mitochondria, and has been reported to be associated with the outer mitochondrial membrane in colon cancer cells, and stimulates mitochondrial bioenergetics (R14)

Enhancing SIRT1 and SIRT3

H2S has been reported to increase NAD+ levels in the vascular endothelium and H₂S itself, associated sulfhydration and NAD+ are decreased during aging. The sirtuins, SIRT1 and SIRT3 are sulfhydrated, which enhances their activity. Accordingly boosting NAD+ levels may improve overall health and lifespan. (R14)

Hypoxia

H₂S produced during normoxic conditions is oxidized in the mitochondria, while during hypoxia this degradation is decreased, leading to an increase in its levels. … it was shown that hypoxia increases mitochondrial H₂S in cardiomyocytes, suggesting a role for the gasotransmitter in oxygen sensing.

Interestingly, concentration of cysteine, the substrate for generation of H₂S, was reported to be about three-fold higher in the mitochondria. The same study also reported mitochondrial translocation of CSE during hypoxia.

In addition, during hypoxia, mitochondrial CBS pools are no longer targeted for degradation by the Lon protease due to deoxygenation of its heme group, leading to a six-fold increase in the CBS.

mTORC1-Sch9 regulates H₂S production via transsulfuration pathway when Cysteine is low

Together, these data indicate that the decreased H₂S production by mTORC1-Sch9 inhibition is most likely due to an increase in the level of intracellular cysteine. … Increased intracellular cysteine usually inhibits the expression of enzymes in the transsulfuration pathway that required for H2S production.

To investigate if the expression of the transsulfuration enzymes is altered in Δsch9 cell, the mRNA levels of CYS3 and CYS4 which encodes Cystathionine gamma-lyase and Cystathionine beta-synthase respectively in yeast were compared in Δsch9 and WT cells.

Indeed, the mRNA levels of both CYS3 and CYS4 decreased to about 50% in Δsch9 cell compared to them in WT cell. And it can be reversed by adding SCH9 back to mutant cells (Figure 5A).

Similarly, inhibiting mTORC1-Sch9 by rapamycin also decreased the expression of both CYS3 and CYS4 (Figure 5B). These data suggest that inhibiting mTORC1-Sch9 which increases intracellular cysteine level does down-regulate transsulfuration pathway. (R15)

The same study confirms this effect in human cells:

The supplementation of Cys/PLP decreased the cell viability, but significantly increased H₂S production. With these assay conditions rapamycin treated cells have decreased the cell death induced by Cys/PLP and higher cell density.

However, less H₂S is produced in both 293T and HeLa cells with rapamycin treatments (Figure 6A and 6B), similar to what we observed in yeast cells (Figure 2D).

These data suggest that inhibiting mTORC1 in mammalian cells may also decrease H₂S production. (R15)

H2S blocks mTOR signalling and induces autophagy

GYY4137 is a novel H2S donor that is stable in vivo and in vitro. …  In this study, we found that GYY4137 could alleviate septicemia-induced ferroptosis in ALI by increasing the expression of GPx4 and SLC7A11 in lung tissue after CLP.    One unexpected finding was the extent to which the levels of ferritin and ferritin light chain increased after CLP, which may be a compensatory mechanism for storing abnormally increased iron.    We also found that the expression of p-mTOR, P62, and Beclin1 was significantly increased and that LC3II/LC3I declined after LPS stimulation, but the effect was inhibited by treatment with GYY4137, indicating that GYY4137 could stimulate (correction in R) the activation of autophagy in sepsis-induced ALI by blocking mTOR signaling. (R21)

Inactivates PTP1B

we report that PTP1B, the founding member of this enzyme family, was reversibly inactivated by H₂S, in vitro and in vivo, via sulfhydration of the active site Cys residue.

Unlike oxidized PTP1B, the sulfhydrated enzyme was preferentially reduced by thioredoxin in vitro, compared to glutathione or dithiothreitol.

Sulfhydration of the active site Cys in PTP1B in cells required the presence of cystathionine-γ-lyase (CSE), a critical enzyme in H₂S production, and resulted in inhibition of phosphatase activity (R16)

Interaction with Iron homeostasis

We further found that H₂S induced more aconitase activity of iron regulatory protein 1 (IRP1) but inhibited its RNA binding activity accompanied with increased protein levels of ferritin and ferriportin, which would contribute to the lower level of labile iron level inside the cells.

In addition, iron was able to suppress CSE-derived H₂S generation, while iron also non-enzymatically induced H₂S release from cysteine. (R17)

High amount of H2S induces Oxidative Stress

Hydrogen sulfide (H2S) is readily water soluble, and, at physiological pH, about two-thirds exists as hydrogen sulfide ion (HS—) and one-third as undissociated H2S.

We use the generic term “sulfide” to refer to both species. Sulfide is an endogenous signal transmitter via protein sulfhydration, and, at low intracellular concentrations—0.01 to 1 μM—donates electrons to complex II of the mitochondrial electron transport chain, thereby stimulating ATP production.

At about 3–30-fold higher concentrations, sulfide becomes toxic by binding to and inhibiting cytochrome C oxidase in complex IV of the electron transport chain, the last complex in the chain prior to ATP synthesis by complex V.

Cyanide, which also inhibits cytochrome C oxidase, increases mitochondrial generation of superoxide and induces oxidative stress in cells (R18)

Detox of some heavy metals, but blocks copper export

Recently, these molecules (reactive sulfide species) were demonstrated to be involved in the detoxification of heavy metals such as methylmercury and zinc.

On the contrary, the current study demonstrated that the combination of H₂S and CuSO4 elicits marked intracellular Cu accumulation, in part, via decreased protein levels of the Cu exporter ATP7A, and leads to increased Cu-dependent neurotoxicity. These findings suggest that H₂S serves as a regulator of the cellular dynamics of heavy metals. (R19)

although methylmercury (MeHg), an environmental toxin, is known to cause neuronal cell death, H₂S and RSS were reported to react with MeHg to form the detoxified metabolite bismethylmercury sulfide ((MeHg)₂S) and protect against MeHg-induced neurotoxicity (R19)