Effects of phenanthridine derivatives on melanogenic protein in
B16 cells
According to the regulatory networks underlying melanogenesis,
upregulation of Wnt/β -catenin signalling promotes the expression
of a critical melanogenic protein, melanocyte-inducing transcription
factor (MITF), which subsequently activates TYRP (tyrosinase-related
protein) 1 and 2 and TYR (tyrosinase) genes and finally increases
melanogenesis (Fig. 1d) (Liu et al., 2019; Manga, Sheyn, Yang,
Sarangarajan & Boissy, 2006). MITF controls the production of the
pigment melanin, and TYRP1 and 2 are two transmembrane proteins. TYRP-1
has been reported to influence TYR activity via forming a complex with
it and/or stabilizing it. TRP-2 functions as a dopachrome tautomerase
downstream of TYR in the melanogenic pathway and regulates the quantity
and quality of melanin produced during melanin biosynthesis. (Wang et
al., 2017). As phenanthridine derivatives could significantly increase
melanin synthesis, it is necessary to further explore whether and how
phenanthridine derivatives affect the expression of MITF, TYRP 1 and 2,
and TYR (tyrosinase). As shown in Fig. 1e, after 48 h of treatment with
HCJA121 and HCJA404 (20 μ M), the MITF gene levels were
significantly enhanced in the treatment groups. Simultaneously, the
levels of the TYRP-1 , TYRP-2 , and TYR genes were
significantly increased in the administration groups . Particularly,
HCJA121 treatment resulted in the strongest activation of MITF, TYRP-1,
and TYR, the levels of which were several-fold higher than those of the
control. To further elucidate the effect of HCJA121 on the expression of
melanogenic proteins (MITF TYRP-1, MITF TYRP-2, and MITF TYR) at the
protein level, western blotting was performed using lysates of B16
murine melanoma cells that were treated with different doses of HCJA121.
As presented in Fig. 1f, compared to the control treatment, HCJA121
treatment clearly increased the expression of MITF TRP-1 and TRP-2 by in
a dose-dependent manner in the B16 cells. These results indicated that
HCJA121 and HCJA404 could simultaneously upregulate all key melanogenic
proteins, which suggested that these compounds may be upstream
regulators of the MITF-TYRP/TYR melanogenic pathway.