Molecular attributes and apoptosis-inducing activities of a putative serine protease isolated from Tiger Milk mushroom (Lignosus rhinocerus) sclerotium against breast cancer cellsin vitro

Abstract

BackgroundThe highly valued medicinal tiger milk mushroom (also known as Lignosus rhinocerus) has the ability to cure numerous ailments. Its anticancer activities are well explored, and recently a partially purified cytotoxic protein fraction termed F5 from the mushroom’s sclerotial cold water extract consisting mainly of fungal serine proteases was found to exhibit potent selective cytotoxicity against a human breast adenocarcinoma cell line (MCF7) with IC50 value of 3.00 μg/ml. However, characterization of its cell death-inducing activity has yet to be established.MethodsThe mechanism involved in the cytotoxic activities of F5 against MCF7 cells was elucidated by flow cytometry-based apoptosis detection, caspases activity measurement, and expression profiling of apoptosis markers by western blotting. Molecular attributes of F5 were further mined from L. rhinocerus’s published genome and transcriptome for future exploration.Results and DiscussionApoptosis induction in MCF7 cells by F5 may involve a cross-talk between the extrinsic and intrinsic apoptotic pathways with upregulation of caspase-8 and -9 activities and a marked decrease of Bcl-2. On the other hand, the levels of pro-apoptotic Bax, BID, and cleaved BID were increased accompanied by observable actin cleavage. At gene level, F5 composed of three predicted non-synonymous single nucleotide polymorphisms (T > C) and an alternative 5′ splice site.ConclusionsFindings from this study provide an advanced framework for further investigations on cancer therapeutics development from L. rhinocerus.

HIV-1 protease and reverse transcriptase inhibition by tiger milk mushroom (Lignosus rhinocerus) sclerotium extracts: In vitro and in silico studies

“…Lignosus rhinocerus (LR), known as the tiger milk mushroom, is traditionally used as folk medicine in Southeast Asia and China. The medicinally beneficial part of LR is the sclerotium, an underground hardened part of the mushroom 1 that has been reported to have several medicinal properties such as neurostimulation, 2 immunomodulation, anti-inflammation, 3 anti-oxidation, anti-proliferation, 4 , 5 anti-diabetes 6 and especially antiviral activity. 7 …”

Proteome Analysis and In Vitro Antiviral, Anticancer and Antioxidant Capacities of the Aqueous Extracts of Lentinula edodes and Pleurotus ostreatus Edible Mushrooms

“…Serine proteinase contributes to the antiviral activity of edible mushrooms [ 30 , 31 ]. Moreover, Yap et al (2018) reported a strong selective cytotoxicity of serine proteinase in the face of a human breast adenocarcinoma cell line (MCF7) and suggested that the mechanism involves the collaborative effect of both extrinsic and intrinsic cell death mechanisms, in addition to the stimulation of caspase-8 and -9 and inhibition of Bcl-2 [ 41 ].…”

Proteins from Lignosus tigris with selective apoptotic cytotoxicity towards MCF7 cell line and suppresses MCF7-xenograft tumor growth

“…MCF7 cells were treated with HMWp at IC 50 value for 24 h, 48 h and 72 h. Total protein was extracted from the treated cells by using RIPA lysis buffer as described by Yap et al (2018) . Thirty micrograms of protein were separated on 12.5% SDS-PAGE and transferred onto polyvinylidene difluoride membrane.…”

“…The cold water extract prepared from sclerotium of L. rhinocerus has also been reported to exhibit cytotoxicity against of a panel of human adenocarcinoma and carcinoma cell lines including breast, lung, leukemic, liver, colorectal, prostate, oral squamous and nasopharyngeal cancers, and proteins and polysaccharide-protein complexes have been hypothesized as the active cytotoxic components ( Lee et al, 2012 ; Lau et al, 2013 ). A subtilisin-like serine protease fraction purified from L. rhinocerus sclerotia was also shown to exhibit cytotoxicity against MCF7 cells via induction of apoptosis ( Yap et al, 2018 ).…”