title: Microinjection of specific anti-IMPDH2 antibodies induces disassembly of cytoplasmic rods/rings that are primarily stationary and stable structures.
abstract: BACKGROUND
  Our laboratory previously reported interesting rods 3-10 μm long and rings 2-5 μm diameter (RR) in the cytoplasm of mammalian cells. Experimental evidence show that both inosine-5'-monophosphate dehydrogenase 2 (IMPDH2) and cytidine triphosphate synthetase (CTPS) are components of RR structures. Several cell types, including mouse embryonic stem cells, and cell lines, such as mouse 3 T3 and rat NRK, naturally present RR structures, while other cells can present RR when treated with compounds interfering with GTP/CTP biosynthetic pathways. In this study, we aimed to investigate the dynamic behavior of these RR in live cells.
  
  
  RESULTS
  RR were detected in >90% of COS-7 and HeLa cells treated with 1 mM ribavirin or 6-Diazo-5-oxo-L-norleucine (DON) for 24 h, and in 75% of COS-7 cells treated with 1 mM mycophenolic acid (MPA) for the same period of time. Microinjection of affinity-purified anti-IMPDH2 antibodies in live COS-7 cells treated with ribavirin, DON, or MPA showed mature forms of RR presented as stable and stationary structures in 71% of cells. In the remaining 29% of cells, RR acquired erratic movement and progressively disassembled into fragments and disappeared within 10 min. The specific stationary state and antibody-dependent disassembling of RR structures was independently confirmed in COS-7 and HeLa cells transfected with GFP-tagged IMPDH2.
  
  
  CONCLUSIONS
  This is the first demonstration of disassembly of RR structures upon microinjection of anti-IMPDH2 antibodies that led to the disappearance of the molecular aggregates. The disassembly of RR after microinjection of anti-IMPDH2 antibody further strengthens the notion that IMPDH2 are major building blocks of RR. Using two independent methods, this study demonstrated that the induced RR are primarily stationary structures in live cells and that IMPDH2 is a key component of RR.
authors.affiliation: Department of Oral Biology, University of Florida, 1395 Center Drive, Gainesville, FL 32610-0424 USA ; Rheumatology Division, Universidade Federal de São Paulo, Rua Botucatu 740, São Paulo, SP 04023-062 Brazil.
authors.forename: Gerson Dierley
authors.initials: GD
authors.lastname: Keppeke
authors.affiliation: Rheumatology Division, Universidade Federal de São Paulo, Rua Botucatu 740, São Paulo, SP 04023-062 Brazil ; Immunology Division, Fleury Medicine and Health Laboratories, Avenida Gal Waldomiro Lima 508, São Paulo, SP 04102-050 Brazil.
authors.forename: Luís Eduardo C
authors.initials: LE
authors.lastname: Andrade
authors.affiliation: Department of Biological Sciences, University of Idaho, 875 Center Drive, Moscow, ID 83844 USA.
authors.forename: Scott S
authors.initials: SS
authors.lastname: Grieshaber
authors.affiliation: Department of Oral Biology, University of Florida, 1395 Center Drive, Gainesville, FL 32610-0424 USA.
authors.forename: Edward K L
authors.initials: EK
authors.lastname: Chan
keywords: CTP synthase; Cytoophidium; Inosine monophosphate dehydrogenase; Intracellular compartment; Mycophenolic acid; Ribavirin