Microinjection of ADP-ribosylated actin inhibits actin synthesis in hepatocyte-hepatoma hybrid cells

• Published in: Biochemical journal 1996-11, Vol.319 ( Pt 3) (3), p.843-849
• Main Authors: Reuner, K H, van der Does, A, Dunker, P, Just, I, Aktories, K, Katz, N
• Format: Article
• Language: English
• Subjects: Actins - biosynthesis; Actins - isolation & purification; Actins - pharmacology; Adenosine Diphosphate Ribose - metabolism; Animals; Botulinum Toxins - pharmacology; Cells, Cultured; Homeostasis; Hybrid Cells - drug effects; Hybrid Cells - metabolism; Liver - metabolism; Liver Neoplasms, Experimental - metabolism; Phalloidine - pharmacology; Rats; Rats, Wistar; RNA, Messenger - analysis; RNA, Messenger - biosynthesis; Serum Albumin - biosynthesis; Transcription, Genetic
• ISSN: 0264-6021; 1470-8728
• DOI: 10.1042/bj3190843

Treatment of hepatocyte-hepatoma hybrid cells with Clostridium botulinum C2 toxin led to a 167% increase in monomeric globular actin (G-actin) and to a 57% decrease in filamentous actin (F-actin) within 2 h. Simultaneously, the level of actin mRNA was specifically decreased to 49% and actin synthesis was significantly diminished. In contrast, treatment of hybrid cells with phalloidin led to a decrease in G-actin to 55% and to a reciprocal increase in actin mRNA to 244% and an increase in actin synthesis. These alterations of actin synthesis depending on the G-actin/F-actin ratio corresponded to the autoregulation of actin synthesis observed in primary cultures of rat hepatocytes. Microinjection of C2 toxin or of phalloidin into hepatocyte-hepatoma hybrid cells had the same effects on actin synthesis as incubation with either toxin in the culture medium. Microinjection of nonpolymerizable ADP-ribosylated G-actin into hepatocyte-hepatoma hybrid cells specifically decreased the incorporation of [35S]methionine into newly synthesized actin within 1 h. This decrease continued for at least 19 h. Microinjection of ADP-ribosylated actin led to rounding of cells and obvious disaggregation of actin filaments, which might be due to capping of actin filaments by the ADP-ribosylated actin. Because stabilization of actin filaments by phalloidin before microinjection of ADP-ribosylated actin also resulted in decreased actin synthesis, the concentration of monomeric G-actin seems to be responsible for the regulation of actin synthesis in hepatocyte-hepatoma hybrid cells, which can be regarded as immortalized hepatocytes.