and
are the causal agents of whooping cough in humans. They produce diverse virulence factors, including adenylate cyclase-hemolysin (AC-Hly), a secreted toxin of the repeat in toxins (RTX) family ...with cyclase, pore-forming, and hemolytic activities. Post-translational modifications (PTMs) are essential for the biological activities of the toxin produced by
. In this study, we compared AC-Hly toxins from various clinical isolates of
and
, focusing on (i) the genomic sequences of
genes, (ii) the PTMs of partially purified AC-Hly, and (iii) the cytotoxic activity of the various AC-Hly toxins. The genes encoding the AC-Hly toxins of
and
displayed very limited polymorphism in each species. Most of the sequence differences between the two species were found in the C-terminal part of the protein. Both toxins harbored PTMs, mostly corresponding to palmitoylations of the lysine 860 residue and palmoylations and myristoylations of lysine 983 for
and AC-Hly and palmitoylations of lysine 894 and myristoylations of lysine 1017 for
AC-Hly. Purified AC-Hly from
was cytotoxic to macrophages, whereas that from
was not.
Pituitary adenylate cyclase-activating polypeptide (PACAP) has been demonstrated to play a crucial part in protecting retinal ganglion cells (RGCs) from apoptosis in various retinal injury animal ...models. PACAP has two basic groups of receptors: PACAP receptor type 1 (PAC1R) and vasoactive intestinal polypeptide/PACAP receptors (VPAC1R and VPAC2R). However, few studies illustrated the spatial and temporal expression changes of endogenous PACAP and its receptors in a rodent optic nerve crush (ONC) model. In this study, a significant upregulation of PACAP and PAC1R in the retina after ONC was observed in both protein and RNA levels. The peak level of PACAP and PAC1R expression could be found on the fifth day following ONC. In addition, immunofluorescent labeling indicated that PACAP and PAC1R were localized mainly in RGCs. On the contrary, VPAC1R and VPAC2R were hardly detected in the retina. Collectively, the spatiotemporal expression of PACAP and its high-affinity receptor PAC1R were remarkably changed after ONC, and mainly expressed in the ganglion cell layer of the retina. This suggested that the upregulation of PACAP and PAC1R may play a vital role in RGC death after ONC.
The RTX (repeats-in-toxin) domain of the bacterial toxin adenylate cyclase (CyaA) contains five RTX blocks (RTX-i to RTX-v) and its folding is essential for CyaA's functions. It was shown that the ...C-terminal capping structure of RTX-v is critical for the whole RTX to fold. However, it is unknown how the folding signal transmits within the RTX domain. Here we use optical tweezers to investigate the interplay between the folding of RTX-iv and RTX-v. Our results show that RTX-iv alone is disordered, but folds into a Ca
-loaded-β-roll structure in the presence of a folded RTX-v. Folding trajectories of RTX-iv-v reveal that the folding of RTX-iv is strictly conditional upon the folding of RTX-v, suggesting that the folding of RTX-iv is templated by RTX-v. This templating effect allows RTX-iv to fold rapidly, and provides significant mutual stabilization. Our study reveals a possible mechanism for transmitting the folding signal within the RTX domain.
Exposure to traumatic stress can lead to fear dysregulation, which has been associated with posttraumatic stress disorder (PTSD). Previous work showed that a polymorphism in the PACAP-PAC1R ...(pituitary adenylate cyclase-activating polypeptide) system is associated with PTSD risk in women, and PACAP (ADCYAP1)-PAC1R (ADCYAP1R1) are highly expressed in the hypothalamus. Here, we show that female mice subjected to acute stress immobilization (IMO) have fear extinction impairments related to Adcyap1 and Adcyap1r1 mRNA upregulation in the hypothalamus, PACAP-c-Fos downregulation in the Medial Amygdala (MeA), and PACAP-FosB/ΔFosB upregulation in the Ventromedial Hypothalamus dorsomedial part (VMHdm). DREADD-mediated inhibition of MeA neurons projecting to the VMHdm during IMO rescues both PACAP upregulation in VMHdm and the fear extinction impairment. We also found that women with the risk genotype of ADCYAP1R1 rs2267735 polymorphism have impaired fear extinction.
In the subgranular zone (SGZ) of the hippocampus, neurogenesis persists throughout life and is upregulated following ischemia. Accumulating evidence suggests that enhanced neurogenesis stimulated by ...ischemic injury contributes to recovery after stroke. However, the mechanisms underlying the upregulation of neurogenesis are unclear. We have demonstrated that a neuropeptide, pituitary adenylate cyclase-activating polypeptide (PACAP), exerts a wide range of effects on neural stem cells (NSCs) during neural development. Here, we examined the effects of endogenous and exogenous PACAP in adult NSCs of the SGZ. Immunostaining showed expression of the PACAP receptor PAC1R in nestin-positive NSCs of adult naive mice. PACAP injection into the lateral ventricle increased bromodeoxyuridine (BrdU)-positive proliferative cells in the SGZ. These data suggest that PACAP promoted the proliferation of NSCs. In global ischemia model mice, the number of BrdU-positive cells was increased in wild-type mice but not in PACAP heterozygous knockout mice. The BrdU-positive cells that increased in number after ischemia were immunopositive for SOX2, a marker of NSCs, and differentiated into NeuN-positive mature neurons at 4 weeks after ischemia. These findings suggest that PACAP contributes to the proliferation of NSCs and may be associated with recovery after brain injury.
We have recently found higher circulating levels of pituitary adenylate cyclase-activating polypeptide (PACAP) associated with posttraumatic stress disorder (PTSD) symptoms in a highly traumatized ...cohort of women but not men. Furthermore, a single nucleotide polymorphism in the PACAP receptor gene ADCYAP1R1 , adenylate cyclase activating polypeptide 1 receptor type 1, was associated with individual differences in PTSD symptoms and psychophysiological markers of fear and anxiety. The current study outlines an investigation of individual differences in brain function associated with ADCYAP1R1 genotype. Forty-nine women who had experienced moderate to high levels of lifetime trauma participated in a functional MRI task involving passive viewing of threatening and neutral face stimuli. Analyses focused on the amygdala and hippocampus, regions that play central roles in the pathophysiology of PTSD and are known to have high densities of PACAP receptors. The risk genotype was associated with increased reactivity of the amygdala and hippocampus to threat stimuli and decreased functional connectivity between the amygdala and hippocampus. The findings indicate that the PACAP system modulates medial temporal lobe function in humans. Individual differences in ADCYAP1R1 genotype may contribute to dysregulated fear circuitry known to play a central role in PTSD and other anxiety disorders.
Summary
Background
Sweat secretion is the major function of eccrine sweat glands; when this process is disturbed (paridrosis), serious skin problems can arise. To elucidate the causes of paridrosis, ...an improved understanding of the regulation, mechanisms and factors underlying sweat production is required. Pituitary adenylate cyclase‐activating polypeptide (PACAP) exhibits pleiotropic functions that are mediated via its receptors PACAP‐specific receptor (PAC1R), vasoactive intestinal peptide (VIP) receptor type 1 (VPAC1R) and VPAC2R. Although some studies have suggested a role for PACAP in the skin and several exocrine glands, the effects of PACAP on the process of eccrine sweat secretion have not been examined.
Objectives
To investigate the effect of PACAP on eccrine sweat secretion.
Methods
Reverse transcriptase‐polymerase chain reaction and immunostaining were used to determine the expression and localization of PACAP and its receptors in mouse and human eccrine sweat glands. We injected PACAP subcutaneously into the footpads of mice and used the starch–iodine test to visualize sweat‐secreting glands.
Results
Immunostaining showed PACAP and PAC1R expression by secretory cells from mouse and human sweat glands. PACAP immunoreactivity was also localized in nerve fibres around eccrine sweat glands. PACAP significantly promoted sweat secretion at the injection site, and this could be blocked by the PAC1R‐antagonist PACAP6‐38. VIP, an agonist of VPAC1R and VPAC2R, failed to induce sweat secretion.
Conclusions
This is the first report demonstrating that PACAP may play a crucial role in sweat secretion via its action on PAC1R located in eccrine sweat glands. The mechanisms underlying the role of PACAP in sweat secretion may provide new therapeutic options to combat sweating disorders.
What's already known about this topic?
Pituitary adenylate cyclase‐activating polypeptide (PACAP) exhibits pleiotropic functions in the central nervous system, including neurotransmission, neuroprotection and vasodilatation.
In the skin, PACAP and its receptors were reported to be important mediators of cutaneous vasoregulation and neurogenic inflammation; however, their involvement in eccrine sweat secretion has not been examined.
What does this study add?
Subcutaneous administration of PACAP into the mouse footpad was found to promote sweat secretion.
This effect was mediated through PAC1R expressed in the secretory cells of eccrine sweat glands.
Most PAC1R‐immunopositivity was observed in these secretory cells.
Immunoreactivity to PACAP was observed in nerve fibres around sweat glands and in the secretory cells.
These findings suggest that PACAP may provide new therapeutic options to modulate the sweating response.
What is the translational message?
Our findings suggest new mechanisms of sweat secretion underlying the role of PACAP.
Further investigation to elucidate detailed molecular mechanisms may offer new perspectives to combat sweating disorders involving hyperhidrosis and anhidrosis, such as Sjögren syndrome, Fabry disease, atopic dermatitis, lichen planus and psoriasis.
Respond to this article
Linked Comment: Cui and Schlessinger. Br J Dermatol 2017; 176:295–296.
PAC1-R is a recognized preferential receptor for the neuropeptide of pituitary adenylate cyclase-activating polypeptide (PACAP), which mediates neuroprotective and nerve regenerative activities of ...PACAP. In this study, we found that in both PAC1R-CHO cells with high expression of PAC1R-eGFP and retinal ganglion cells (RGC-5) with the natural expression of PAC1-R, oligo-peptide PACAP(28-38) and the positively charged arginine-rich penetrating peptide TAT, as positive allosteric modulators of PAC1-R, significantly trigger the nuclear translocation of PAC1-R. The chromatin immunoprecipitation (ChIP)-PCR results show that the nuclear translocated PAC1-R binds with the promoter regions of
and its specific ligand
. The up-regulated promoter activities of
and
induced by PACAP(28-38) or TAT are positively correlative with the increase of the expression levels of PAC1-R and PACAP. Moreover, the nuclear translocation of PAC1-R induced by PACAP(28-38) or TAT is significantly inhibited by the mutation of PAC1-R on Cys25 and the palmitoylation inhibitor 2-bromopalmitate. Meanwhile, the increase in both PAC1-R and PACAP levels and the neuroprotective activities of PACAP(28-38) and TAT in MPP-induced cell model of Parkinson ' s disease are synchronously inhibited by 2-bromopalmitate, which are positively correlated with the nuclear translocation of PAC1-R induced by PACAP(28-38) or TAT. Bioinformatics analysis and motif enrichment analysis following ChIP-sequencing show that the transcription factors including SP1, Zic2, GATA1, REST and YY1 may be recruited by nuclear PAC1-R and involved in regulating the promoter activities of
and
. ChIP-sequencing and related bioinformatics analysis show that the downstream target genes regulated by the nuclear PAC1-R are mostly involved in the process of cellular stress and related to neuroprotection, neuronal genesis and development.
Mice lacking pituitary adenylate cyclase-activating polypeptide (PACAP) display psychomotor abnormalities, most of which are ameliorated by atypical antipsychotics with serotonin (5-HT) 2A receptor ...(5-HT
) antagonism. Heterozygous
mutant mice show a significantly higher hallucinogenic response than wild-type mice to a 5-HT
agonist. Endogenous PACAP may, therefore, affect 5-HT
signaling; however, the underlying neurobiological mechanism for this remains unclear. Here, we examined whether PACAP modulates 5-HT
signaling by addressing cellular protein localization. PACAP induced an increase in internalization of 5-HT
but not 5-HT
, 5-HT
, dopamine D
receptors or metabotropic glutamate receptor 2 in HEK293T cells. This PACAP action was inhibited by protein kinase C inhibitors, β-arrestin2 silencing, the PACAP receptor PAC1 antagonist PACAP
, and PAC1 silencing. In addition, the levels of endogenous 5-HT
were decreased on the cell surface of primary cultured cortical neurons after PACAP stimulation and were increased in frontal cortex cell membranes of
mice. Finally, intracerebroventricular PACAP administration suppressed 5-HT
agonist-induced head twitch responses in mice. These results suggest that PACAP-PAC1 signaling increases 5-HT
internalization resulting in attenuation of 5-HT
-mediated signaling, although further study is necessary to determine the relationship between behavioral abnormalities in
mice and PACAP-induced 5-HT
internalization.
Neuropeptides may exert trophic effects during development, and then neurotransmitter roles in the developed nervous system. One way to associate peptide-deficiency phenotypes with either role is ...first to assess potential phenotypes in so-called constitutive knockout mice, and then proceed to specify, regionally and temporally, where and when neuropeptide expression is required to prevent these phenotypes. We have previously demonstrated that the well-known constellation of behavioral and metabolic phenotypes associated with constitutive pituitary adenylate cyclase-activating peptide (PACAP) knockout mice are accompanied by transcriptomic alterations of two types: those that distinguish the PACAP-null phenotype from wild-type (WT) in otherwise quiescent mice (cPRGs), and gene induction that occurs in response to acute environmental perturbation in WT mice that do not occur in knockout mice (aPRGs). Comparing constitutive PACAP knockout mice to a variety of temporally and regionally specific PACAP knockouts, we show that the prominent hyperlocomotor phenotype is a consequence of early loss of PACAP expression, is associated with Fos overexpression in hippocampus and basal ganglia, and that a thermoregulatory effect previously shown to be mediated by PACAP-expressing neurons of medial preoptic hypothalamus is independent of PACAP expression in those neurons in adult mice. In contrast, PACAP dependence of weight loss/hypophagia triggered by restraint stress, seen in constitutive PACAP knockout mice, is phenocopied in mice in which PACAP is deleted after neuronal differentiation. Our results imply that PACAP has a prominent role as a trophic factor early in development determining global central nervous system characteristics, and in addition a second, discrete set of functions as a neurotransmitter in the fully developed nervous system that support physiological and psychological responses to stress.
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