Marine-derived microbes and molecules for drug discovery.

Increasing attention has been paid to marine-derived biomolecules as sources of therapeutics for autoimmune diseases. Nagasaki Prefecture has many islands and is surrounded by seas, straits, gulfs, bays, and coves, giving it the second longest coastline in Japan after Hokkaido. We have collected more than 20,000 marine microbes and have been preparing an original marine microbial extract library, which contains small and mid-size biomolecules that may penetrate cell membranes and interfere with the intracellular protein-protein interaction involved in the development of autoinflammatory diseases such as familial Mediterranean fever. In addition, we have been developing an indoor shark farming system to prepare shark nanobodies that could be developed as potential therapeutic agents for autoimmune diseases. Sharks produce heavy-chain antibodies, called immunoglobulin new antigen receptors (IgNARs), consisting of one variable domain (VNAR) and five constant domains (CNAR); of these, VNAR can recognize a variety of foreign antigens. A VNAR single domain fragment, called a nanobody, can be expressed in Escherichia coli and has the properties of an ideal therapeutic candidate for autoimmune diseases. Shark nanobodies contain complementarity-determining regions that are formed through the somatic rearrangement of variable, diversity, and joining segments, with the segment end trimming and the N- and P-additions, as found in the variable domains of mammalian antibodies. The affinity and diversity of shark nanobodies are thus expected to be comparable to those of mammalian antibodies. In addition, shark nanobodies are physically robust and can be prepared inexpensively; as such, they may lead to the development of highly specific, stable, effective, and inexpensive biotherapeutics in the future. In this review, we first summarize the history of the development of conventional small molecule drugs and monoclonal antibody therapeutics for autoimmune diseases, and then introduce our drug discovery system at Nagasaki University, including the preparation of an original marine microbial extract library and the development of shark nanobodies.

Cleaved PGAM5 dephosphorylates nuclear serine/arginine-rich proteins during mitophagy.

PGAM5 is a protein phosphatase located in the inner mitochondrial membrane through its transmembrane (TM) domain and is cleaved within the TM domain upon mitochondrial dysfunction. We found previously that cleaved PGAM5 is released from mitochondria, following proteasome-mediated rupture of the outer mitochondrial membrane during mitophagy, a selective form of autophagy specific to mitochondria. Here, we examined the role of cleaved PGAM5 outside mitochondria. Deletion mutants that mimic cleaved PGAM5 existed not only in the cytosol but also in the nucleus, and a fraction of cleaved PGAM5 translocated to the nucleus during mitophagy induced by the uncoupler CCCP. We identified serine/arginine-related nuclear matrix protein of 160 kDa (SRm160)/SRRM1, which contains a highly phosphorylated domain rich in arginine/serine dipeptides, called the RS domain, as a nuclear protein that interacts with PGAM5. PGAM5 dephosphorylated SRm160, and incubation of lysates from WT cells, but not of those from PGAM5-deficient cells, induced dephosphorylation of SRm160 and another RS domain-containing protein SRSF1, one of the most characterized serine/arginine-rich (SR) proteins. Moreover, phosphorylation of these proteins and other SR proteins, which are commonly reactive toward the 1H4 monoclonal antibody that detects phosphorylated SR proteins, decreased during mitophagy, largely because of PGAM5 activity. These results suggest that PGAM5 regulates phosphorylation of these nuclear proteins during mitophagy. Because SRm160 and SR proteins play critical roles in mRNA metabolism, PGAM5 may coordinate cellular responses to mitochondrial stress at least in part through post-transcriptional and pre-translational events.