When Cytokinin, a Plant Hormone, Meets the Adenosine A2A Receptor: A Novel Neuroprotectant and Lead for Treating Neurodegenerative Disorders?

It is well known that cytokinins are a class of phytohormones that promote cell division in plant roots and shoots. However, their targets, biological functions, and implications in mammalian systems have rarely been examined. In this study, we show that one cytokinin, zeatin riboside, can prevent pheochromocytoma (PC12) cells from serum deprivation-induced apoptosis by acting on the adenosine A2A receptor (A2A-R), which was blocked by an A2A-R antagonist and a protein kinase A (PKA) inhibitor, demonstrating the functional ability of zeatin riboside by mediating through A2A-R signaling event. Since the A2A-R was implicated as a therapeutic target in treating Huntington’s disease (HD), a cellular model of HD was applied by transfecting mutant huntingtin in PC12 cells. By using filter retardation assay and confocal microscopy we found that zeatin riboside reversed mutant huntingtin (Htt)-induced protein aggregations and proteasome deactivation through A2A-R signaling. PKA inhibitor blocked zeatin riboside-induced suppression of mutant Htt aggregations. In addition, PKA activated proteasome activity and reduced mutant Htt protein aggregations. However, a proteasome inhibitor blocked both zeatin riboside-and PKA activator-mediated suppression of mutant Htt aggregations, confirming mediation of the A2A-R/PKA/proteasome pathway. Taken together, zeatin riboside might have therapeutic potential as a novel neuroprotectant and a lead for treating neurodegenerative disorders

JC Virus Small t Antigen Binds Phosphatase PP2A and Rb Family Proteins and Is Required for Efficient Viral DNA Replication Activity

BACKGROUND: The human polyomavirus, JC virus (JCV) produces five tumor proteins encoded by transcripts alternatively spliced from one precursor messenger RNA. Significant attention has been given to replication and transforming activities of JCV's large tumor antigen (TAg) and three T' proteins, but little is known about small tumor antigen (tAg) functions. Amino-terminal sequences of tAg overlap with those of the other tumor proteins, but the carboxy half of tAg is unique. These latter sequences are the least conserved among the early coding regions of primate polyomaviruses. METHODOLOGY AND FINDINGS: We investigated the ability of wild type and mutant forms of JCV tAg to interact with cellular proteins involved in regulating cell proliferation and survival. The JCV P99A tAg is mutated at a conserved proline, which in the SV40 tAg is required for efficient interaction with protein phosphatase 2A (PP2A), and the C157A mutant tAg is altered at one of two newly recognized LxCxE motifs. Relative to wild type and C157A tAgs, P99A tAg interacts inefficiently with PP2A in vivo. Unlike SV40 tAg, JCV tAg binds to the Rb family of tumor suppressor proteins. Viral DNAs expressing mutant t proteins replicated less efficiently than did the intact JCV genome. A JCV construct incapable of expressing tAg was replication-incompetent, a defect not complemented in trans using a tAg-expressing vector. CONCLUSIONS: JCV tAg possesses unique properties among the polyomavirus small t proteins. It contributes significantly to viral DNA replication in vivo; a tAg null mutant failed to display detectable DNA replication activity, and a tAg substitution mutant, reduced in PP2A binding, was replication-defective. Our observation that JCV tAg binds Rb proteins, indicates all five JCV tumor proteins have the potential to influence cell cycle progression in infected and transformed cells. It remains unclear how these proteins coordinate their unique and overlapping functions

Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)1.

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field