Skin cancer is a worldwide problem and is more common in people with lighter skin than people with darker skin colour. Ultraviolet (UV) radiation is the primary contributor to skin cancer development by causing DNA damage in skin cells called keratinocytes. Melanin, the pigment responsible for skin colour, also plays a role in generating so-called melanin caps – umbrella-like structures that protect the cells’ DNA from UV-related damage. Dr Hongguang Lu and his team from Guizhou Medical University, China, shed light on the cap formation process and identified a photoreceptor called OPN3 as a potential therapeutic target to prevent DNA damage.
Skin cancer is one of the most common malignant tumours worldwide. Epidemiological studies have determined that skin cancer occurs more often in people with lighter skin than people with brown or black skin. Ultraviolet (UV) radiation from the sun is a key contributor to skin cancer development. There are three types of UV – UVA, UVB, and UVC. Both UVA and UVB are carcinogenic; however, UVA compromises most of the UV that reaches the Earth’s surface. UVA affects our DNA in skin cells, causing genetic changes that are responsible for causing cancer.
Melanin is the primary pigment in our body responsible for skin, eye, and hair colour. Our genetics mainly determine the amount of melanin in the body. Melanin also defends us against skin damage, including UV radiation. The pigment is synthesised by melanin-generating cells called melanocytes, and it is commonly transferred into cells called keratinocytes, a typical cell in the outermost layer of skin, the epidermis.
The melanin caps
In 1996, while a guest research fellow at the Department of Dermatology, University Hospital of Wales College of Medicine, UK, Dr Hongguang Lu and his colleagues observed melanin in keratinocytes from people worldwide. They found that melanin forms a cap over the nucleus in keratinocytes, where the DNA and genetic material are contained. These melanin caps reside at the skin surface and protect the keratinocytes from DNA damage caused by UV. Interestingly, these melanin caps are more common in people with darker skin. However, the mechanism behind the formation of the melanin cap is yet to be determined.People with heavily pigmented skin harbour more melanin caps compared to people with lighter skin, and this is associated with lower skin cancer incidence.
The quality and quantity of melanin and melanin caps vary between racial groups. People with heavily pigmented skin harbour more melanin caps compared to people with lighter skin, and this is associated with lower skin cancer incidence. Melanin cap formation is mainly related to skin colour or melanin pigmentation. However, how UV radiation or sunlight can cause a melanin response to form these melanin caps is not well understood in keratinocytes.
The power of light
All living beings on Earth use light photons from the sun as an adaptive advantage; for example, animals and humans use photons to help identify objects through their vision. UV radiation is composed of photons that activate receptors in the eye, called G protein-coupled receptors (GPCRs) and downstream effectors. In most animals, light response pathways use a member of the opsin (OPN) family of GPCRs as a light detector. Over 1,000 OPNs have been identified in the animal kingdom. Some research suggests that these UV-sensing systems may also be present in the skin. One of these opsins, called OPN3, predominantly expressed in the skin, has been found to be important in skin pigmentation regulation and even skin tumour progression. However, it is unclear if OPN3 is involved in the melanin cap formation as a response to UV exposure.
In a recent publication, Lu – now at the Guizhou Medical University in China – and colleagues demonstrated that OPN3 is critical in melanin cap formation and helps to protect the skin from damage like skin cancer. They have also determined the pathway by which OPN3 acts.
Lu and the team found that different doses of UV radiation induce the expression of dynein, a transporter molecule in melanocytes that mediates the transport of melanin to keratinocytes. UV radiation also significantly upregulated proteins called p150Glued (DCTN1) and Cytoplasmic dynein intermediate chains (Dync1i1) in human keratinocytes. These proteins are thought to facilitate the movement of substances within cells.
To confirm whether DCTN1 is involved in UVA-induced keratinocyte melanin cap formation, researchers used small molecules called siRNAs to attach and downregulate DCTN1 levels in keratinocytes.
Given UVA irradiation, the researchers observed that the forming ability of melanin cap reduced in the experiment group, suggesting that DCTN1 is an important factor in forming melanin caps in keratinocytes as a result of UV exposure.
OPN3 expression in keratinocytes
Opsins, particularly OPN3, are believed to play a more significant role in the skin than once thought, due to their high expression in the skin. Research suggests that OPN3 may function as an epidermal photoreceptor in skin keratinocytes and even have other physiological roles.
Lu and his team describe that UVA significantly upregulated OPN3 expression in keratinocytes. Previous studies have described that UVA radiation causes oxidative damage that may induce a melanin response in keratinocytes. However, they demonstrate that 3J/cm2 UVA does not generate reactive oxygen species at a certain range of doses and times, which is responsible for causing oxidative damage. Instead, UVA was found to mediate the expression of Dync1i1 and DCTN1 in keratinocytes through OPN3, which leads to melanin cap formation. The research team also found that blocked OPN3 expression interfered with Dync1i1 and DCTN1 expression induced by UVA. This suggests OPN3 needs to be functional for melanin caps to be formed and that Dync1i1 and DCTN1 function through OPN3. Alternatively, when OPN3 levels were upregulated using virus technology, Dync1i1 and DCTN1 levels also increased in the presence of UVA – confirming that OPN3 is crucial for melanin cap generation through Dync1i1 and DCTN1-related pathways.
Next, Lu and his team wanted to explore what signalling pathways are involved in the formation of the melanin caps, as he previously found that UV drives the movement of calcium stored in melanocytes.
They found that UVA caused calcium inside melanocytes to move via a receptor called transient receptor potential channels, ankyrin 1. UVA also induced the activation of a molecule called CAMKII through phosphorylation (by which a molecule called a phosphate group is added to another molecule to activate or inhibit it). However, they were unsure if the calcium signalling pathway is involved in the formation of melanin caps.
Before UVA exposure, they measured calcium levels inside (intracellular) the keratinocytes. UVA exposure increases intracellular calcium and upregulated activation of CAMKII and CREB. They found that silent OPN3 expression caused the amount of intracellular calcium to decrease in cells when exposed to UVA. Expression of calcium-related proteins was also reduced with the ceased expression of OPN3.
Fitting the pieces together
To decipher whether Dync1i1 and DCTN1 expression were mediated through calcium signalling, calcium signalling pathways were artificially blocked using a PTX substance. PTX blocks G proteins, causing calcium release via activation of a molecule called phospholipase Cβ (PLC). Researchers tested whether the expression of PLCβ was upregulated by UVA exposure. UVA was indeed seen to upregulate PLCβ, but this was not the case when the calcium signalling pathways were blocked. However, UVA exposure caused a decreased expression of PLCβ when OPN3 was inhibited, suggesting that UVA mediates the release of calcium through OPN3.The research paves the way for OPN3 as a potential therapeutic target, protecting humans against UV-related DNA damage and, ultimately, skin cancer.
Researchers further blocked PLCβ in keratinocytes to see if this affected calcium responses. The PLC inhibitor significantly stopped UVA-induced calcium movement. Moreover, UVA exposure increased CAMKII, CREB, Dync1i1 and DCTN1 levels, but this effect ceased when calcium release was inhibited, indicating that UVA mediates the PLCβ/calcium expression through OPN3.
Intracellular calcium movement activates the AKT pathway, which is important in recruiting the necessary proteins required for movement. One protein crucial in this pathway is DCTN1. However, its role in regulating DCTN1 in keratinocytes is unclear. Lu and his team found UVA-induced AKT activation but not when OPN3 expression was silenced or when calcium release was blocked, indicating that UVA regulates the AKT pathway through OPN3/calcium signalling. Interestingly, AKT was important in the regulation of Dync1i1 and DCTN1 regulation, as UVA-induced Dync1i1 and DCTN1 expression was eliminated when AKT activation was blocked.
A better understanding
Lu and his colleague’s findings suggest that UVA-induced DCTN1 and Dync1i1 expression promotes the formation of melanin caps in keratinocytes that is completed via the OPN3/calcium/AKT pathway. This research summarises that OPN3, a UVA photoreceptor, is vital in forming melanin caps in human keratinocytes. These findings expand our knowledge and understanding of the function of OPN3 and its role in the skin as a sensor for UV-induced melanin cap formation in keratinocytes. The research paves the way for OPN3 to be used as a potential therapeutic target, protecting humans against UV-related DNA damage and, ultimately, skin cancer.
Personal ResponseWhat inspired you to conduct this study?
Skin cancer is one of the cancers with the highest incidence rate and mortality in the world, and it has become a major public health problem. Ultraviolet (UV) radiation-induced DNA damage is an important risk factor for the occurrence and development of skin cancer. Melanin cap formation in keratinocytes acts as a natural sunscreen and protects DNA by absorbing and emitting UV radiation. Therefore, our in-depth exploration of the molecular mechanism of UV-induced melanin cap formation is crucial for protecting cellular DNA.
Can this research improve the current treatment of skin cancer?
Some studies have shown that OPN3 may play an important role in the occurrence and development of tumours. Our study provides insights into the molecular mechanisms by which human keratinocytes respond to UVA radiation to induce supranuclear cap formation, which suggests that OPN3 may serve as a therapeutic target for DNA damage caused by ultraviolet radiation. Our team is currently developing inhibitors for OPN3. Of course, a large amount of research is needed to explore the functions of OPN3 in the later stage.