The role of the hippo pathway in melanocytes and melanoma

Abstract

The Hippo signaling pathway comprises a series of cytoplasmic tumor suppressor proteins including Merlin and the Lats1/2 and MST1/2 kinases, and is thought to play a critical role in determining the sizes of organs and tissues. The Hippo pathway is regulated upstream by extracellular mechanosensory signaling arising from cell shape and polarity, as well as by a variety of extracellular signaling molecules. When active, the pathway maintains the transcriptional activators Yes-associated protein (YAP) and TAZ in phosphorylated forms in the cytoplasm, preventing cell proliferation. When the Hippo pathway is inactivated, YAP and TAZ are translocated to the nucleus and induce the expression of a variety of proteins concerned with entry into the cell division cycle, such as cyclin D1 and Fox M1, as well as the inhibition of apoptosis. The failure of the Hippo pathway has been implicated in the development of many different types of cancer but there is limited information available as to its involvement in melanoma. We hypothesize here firstly that the Hippo pathway is involved in maintaining density of cutaneous melanocytes on the basement membrane at the junction of the epidermis and the dermis, and secondly, that its function is disturbed in melanoma. We have analyzed a series of 23 low passage human melanoma lines as well as cultured normal melanoma, and find that melanocytes, as well as all melanoma cell lines examined express TAZ. Melanocytes and most melanoma lines also express YAP. E-cadherin, an upstream regulator of the Hippo pathway, and Axl, a receptor tyrosine kinase regulated by the Hippo pathway, are expressed in melanocytes and in several melanoma cell lines. These observations, together with published evidence for the presence of Merlin, Lats1/2, and MST1/2 in melanocytes and melanoma cells, support the hypothesis that the Hippo pathway is an important component of melanocyte and melanoma behavior.

Keywords: E-cadherin; cell proliferation; cytoskeleton; epidermal melanocytes; merlin.

Figure 1

Diagram of a melanocyte on the basement membrane and sandwiched between the epidermis and the dermis. Melanocytes extend a number of dendrites into the epidermis and these serve to transfer melanosomes from each melanocyte to a number of keratinocytes. Connections between adjacent melanocytes also involve dendrites but are not shown in the figure.

Figure 2

Phase contrast photomicrograph of normal melanocytes growing on a layer of Matrigel.

Figure 3

Diagram showing some of the upstream elements of the proposed Hippo pathway. Melanocytes are strongly polarized and receive signals on the one hand from the epidermis through adherens junctions with keratinocytes, and on the other hand from the basement membrane through integrins. They also receive signals through receptor tyrosine kinases such as Axl and through G-protein coupled receptors (GPCRs). All are likely to signal through kinases to Lats1/2; arrows indicate signaling events.

Figure 4

Diagram showing some of the downstream elements of the proposed Hippo pathway. The transcription factors YAP/TAZ-TEAD are activated, and DYRK-DREAM is inactivated by loss of Hippo signalling. Resulting transcription products are involved in regulating the synthesis of a number of proteins associated with cell proliferation and migration.

Figure 5

Western blots of whole-cell extracts of cultures of normal melanocytes and of a number of melanoma lines, indicating expression of YAP and TAZ. The numbers indicate the identities of members of the New Zealand melanoma collection (e.g., 3 = NZM3). Mel indicates data for normal melanocytes. Data for expression on western blots of E-cadherin and Axl, taken from another publication (Kim et al., submitted), are shown below the NZM numbers; – expression not detected; +weak expression; ++strong expression; nd not done.