Biology subjects

Barral, D. C.

Publications and source records attributed to Barral, D. C..

3 recordsLinked to original sources

bioRxiv2021

Rab3a regulates melanin exocytosis induced by keratinocyte-conditioned medium

Skin pigmentation relies on melanin and is crucial for photoprotection against ultraviolet radiation-induced toxicity. Melanin is synthesized and stored in melanosomes, within melanocytes and then transferred to keratinocytes. While the molecular players involved in melanogenesis have been extensively studied, those underlying melanin transfer remain poorly characterized. Previously, our group proposed that coupled exo/phagocytosis is the predominant mechanism of melanin transfer in human skin and showed an essential role for Rab11b and the exocyst tethering complex in this process. Using a fluorescence-based assay, we show here that keratinocyte-conditioned medium (KCM) specifically induces melanin exocytosis from melanocytes. Moreover, we found that Rab3a, but not Rab11b, regulates melanin exocytosis upon KCM stimulation. In fact, melanosomes accumulate in melanocyte dendrites upon KCM stimulation, co-localizing with Rab3a mainly in the vicinity of the plasma membrane. Additionally, Rab3a silencing does not affect melanin transfer in melanocyte/keratinocyte co-cultures, in contrast with Rab11b depletion, indicating that Rab11b regulates non-KCM-stimulated melanin exocytosis. Thus, our results suggest the existence of at least two distinct routes of melanin exocytosis: one controlled by Rab11b and another Rab3a-dependent, stimulated by KCM. Furthermore, these results provide evidence that soluble factors secreted by keratinocytes can control skin pigmentation via induction of melanocyte signaling pathways that promote peripheral transport of melanosomes and a Rab3a-mediated exocytosis mechanism.

molecular biology↗

bioRxiv2021

Melanocore uptake by keratinocytes occurs through phagocytosis and involves Protease-activated receptor-2 activation

In the skin epidermis, melanin is produced and stored within melanosomes in melanocytes and then transferred to keratinocytes. Different models have been proposed to explain the melanin transfer mechanism, which differ essentially in how melanin is transferred - either in a membrane-bound melanosome or as a melanosome core, i.e. melanocore. Here we investigated the endocytic route followed by melanocores and melanosomes during internalization by keratinocytes, by comparing the uptake of melanocores isolated from the supernatant of melanocyte cultures with melanosomes isolated from melanocytes. We show that inhibition of actin dynamics impairs the uptake of both melanocores and melanosomes. Moreover, depletion of critical proteins involved in actin-dependent uptake mechanisms, namely Rac1 and CtBP1/BARS, together with inhibition of Rac1-dependent signaling pathways or macropinocytosis suggest that melanocores are internalized by phagocytosis, whereas melanosomes are internalized by macropinocytosis. Furthermore, we confirmed that melanocore, but not melanosome uptake is dependent on the Protease-activated receptor-2 (PAR-2) and found that PAR-2 can be specifically activated by melanocores. As skin pigmentation was shown to be regulated by PAR-2 activation, our results further support the melanocore mechanism of melanin transfer and further refine this model, which can now be described as coupled melanocore exo/phagocytosis.

cell biology↗

bioRxiv2021

A novel Rab11-Rab3a cascade required for lysosome exocytosis

Lysosomes are dynamic organelles, capable of undergoing exocytosis. This process is crucial for several cellular functions, namely plasma membrane repair. Nevertheless, the molecular machinery involved in this process is poorly understood. Here, we identify Rab11a and Rab11b as regulators of calcium-induced lysosome exocytosis. Interestingly, Rab11-positive vesicles transiently interact with lysosomes at the cell periphery, indicating that this interaction is required for the last steps of lysosome exocytosis. Additionally, we found that the silencing of the exocyst subunit Sec15, a Rab11 effector, impairs lysosome exocytosis independently of the exocyst complex, suggesting that Sec15 acts together with Rab11 in the regulation of lysosome exocytosis. Furthermore, we show that Rab11 binds the guanine nucleotide exchange factor for Rab3a (GRAB) and also Rab3a, which we described previously as a regulator of the positioning and exocytosis of lysosomes. Thus, our studies suggest that Rab11-positive vesicles transport GRAB to activate Rab3a on lysosomes, establishing a Rab11-Rab3 cascade that is essential for lysosome exocytosis.

cell biology↗