Dataset Information

Opsin 5 is a key regulator of ultraviolet radiation-induced melanogenesis in human epidermal melanocytes.

ABSTRACT:

Background

Human skin, which is constantly exposed to solar ultraviolet radiation (UVR), has a unique ability to respond by increasing its pigmentation in a protective process driven by melanogenesis in human epidermal melanocytes (HEMs). However, the molecular mechanisms used by HEMs to detect and respond to UVR remain unclear.

Objectives

To investigate the function and potential mechanism of opsin 5 (OPN5), a photoreceptor responsive to UVR wavelengths, in melanogenesis in HEMs.

Methods

Melanin content in HEMs was determined using the NaOH method, and activity of tyrosinase (TYR) (a key enzyme in melanin synthesis) was determined by the l-DOPA method. OPN5 expression in UVR-treated vs. untreated HEMs and explant tissues was detected by reverse-transcription quantitative polymerase chain reaction (RT-qPCR), Western blotting and immunofluorescence. Short interfering RNA-mediated OPN5 knockdown and a lentivirus OPN5 overexpression model were used to examine their respective effects on TYR, tyrosinase-related protein 1 (TRP1), TRP2 and microphthalmia-associated transcription factor (MITF) expression, under UVR. Changes in expression of TYR, TRP1 and TRP2 caused by changes in OPN5 expression level were detected by RT-qPCR and Western blot. Furthermore, changes in signalling pathway proteins were assayed.

Results

We found that OPN5 is the key sensor in HEMs responsible for UVR-induced melanogenesis. OPN5-induced melanogenesis required Ca²⁺ -dependent G protein-coupled receptor- and protein kinase C signal transduction, thus contributing to the UVR-induced MITF response to mediate downstream cellular effects, and providing evidence of OPN5 function in mammalian phototransduction. Remarkably, OPN5 activation was necessary for UVR-induced increase in cellular melanin and has an inherent function in melanocyte melanogenesis.

Conclusions

Our results provide insight into the molecular mechanisms of UVR sensing and phototransduction in melanocytes, and may reveal molecular targets for preventing pigmentation or pigment diseases.

SUBMITTER: Lan Y

PROVIDER: S-EPMC8453816 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BackgroundChronic exposure to ultraviolet (UV) radiation (mainly UVA) induces a sustained increase of matrix metalloproteinases (MMPs), especially MMP1, MMP2, MMP3 and MMP9 in human skin fibroblasts. MMPs can lead to the degradation of fibrous connective tissue, which is the main cause of skin photoageing. The molecular mechanisms through which fibroblasts sense UVA and trigger the cell signalling pathways involved in the upregulation of MMPs have not been well elucidated.ObjectivesHere, we investigated the function and potential mechanisms of photoageing of opsin (OPN)3 in normal human dermal fibroblasts (NHDFs).MethodsReal-time polymerase chain reaction and Western blot analysis were used to analyse the expression levels of OPN3 in NHDFs and photoageing models. Subsequently, NHDFs transfected with OPN3 inhibitors and indicators related to photoageing before and after UVA irradiation included expression levels of MMP1, MMP2, MMP3 and MMP9, and signalling pathway protein molecules were examined.ResultsWe provide evidence that OPN3 initiates UVA phototransduction in NHDFs. OPN3 triggers phosphorylation of activator protein 1 and ultimately upregulates MMP1, MMP2, MMP3 and MMP9 in NHDFs through activating Ca2+ /calmodulin-dependent protein kinase II, cyclic adenosine monophosphate response element-binding protein, extracellular signal-regulated kinase, c-JUN N-terminal kinase and p38. Here, we demonstrate for the first time that OPN3 is the key sensor responsible for upregulating MMP1, MMP2, MMP3 and MMP9 in NHDFs following UVA exposure via the calcium-dependent G protein-coupled signalling pathway.ConclusionsOur studies provide insights into the understanding of the molecular mechanisms through which human skin cells respond to UVA radiation and may reveal molecular targets for skin photoageing prevention or clinical management. What's already known about this topic? Photoaged fibroblasts accumulate with long-term ultraviolet (UV) exposure. Matrix metalloproteinases (MMPs) play an important role in the pathogenesis of photoageing. MMP1, MMP2, MMP3 and MMP9 are responsible for the destruction of fibroblast collagen in severely photodamaged skin. Opsins (OPNs) are light-sensitive members of the superfamily of heptahelical G protein-coupled receptors, a family of cell surface receptors that are activated by a variety of stimuli and mediate signalling across membranes. What does this study add? OPN3 is highly expressed in fibroblasts and responds to UVA irradiation. OPN3 regulates the expression of MMP1, MMP2, MMP3 and MMP9 via the calcium-dependent G protein-coupled signalling pathway. OPN3 is a light-sensitive sensor that plays an important role in photoageing of the skin.

Project description:To understand the signalling mechanisms involved in the dual stimulatory effects of endothelin-1 (ET-1) on DNA synthesis and melanization in cultured human melanocytes, we analysed the biological profile of ET-1 receptor and determined the effects of ET-1 on the protein kinase C, cyclic AMP system and mitogen-activated protein kinase (MAP kinase) in comparison with their relevant stimulants. The photoaffinity labelling of ET-1 receptors with Denny-Jaff reagents revealed an ET-1 receptor with a molecular mass of 51 kDa in human melanocytes. The ET(A) receptor subtype-sensitive antagonist BQ123(50 nM) or pertussis toxin (100 ng/ml) significantly suppressed the ET-1-induced intracellular calcium mobilization, indicating the presence of pertussis toxin-sensitive G-protein-coupled ET(A) receptors. An assay of protein kinase C activity revealed that 10nM ET-1 translocated cytosolic protein kinase C to membrane-bound protein kinase C within 5 min of the start of incubation. In contrast, receptor-mediated melanocyte activation by ET-1 was accompanied by an elevated level of cyclic AMP (4-fold over control) after 10-60 min of incubation, whereas 60 min of incubation of human melanocytes with c-Kit or c-Met ligands such as stem cell factor (10 nM) or basic fibroblast growth factor (10 nM) did not elevate the cyclic AMP level. We have also demonstrated that a specific tyrosine kinase inhibitor, tyrphostin B-42 (10 microM), inhibited the ET-1-induced growth stimulation, suggesting the involvement of the tyrosine kinase pathway in growth stimulation. Consistently, an assay of MAP kinase revealed that ET-1 caused a 10-fold activation of MAP kinase after 5 min of incubation with human melanocytes in a similar way to tyrosine kinase ligands such as stem cell factor and hepatocyte growth factor. Further, the DNA synthesis stimulated by the c-Kit ligand stem cell factor at a concentration of 1 nM was synergistically enhanced by 5 nM ET-1. These results suggest that ET-induced dual cellular events in human melanocytes are closely associated with cross-talk between the protein kinase C and A and tyrosine kinase pathways.