TUBULIN POLYGLYCYLATION IN PLATYHELMINTHES - DIVERSITY AMONG STABLE MICROTUBULE NETWORKS AND VERY LATE OCCURRENCE DURING SPERMIOGENESIS

The distribution of glycylated tubulin has been analyzed in different populations of stable microtubules in a digenean flatworm, Echinostoma caproni (Platyhelminthes). Two cellular types, spermatozoa and ciliat ed excretory cells, have been analyzed by means of immunofluorescence, immunogold, and immunoblotting techniques using two monoclonal antibo dies (mAbs), AXO 49, and TAP 952, specifically directed against differ ently glycylated isoforms of tubulin. The presence of glycylated tubul in in the two cell types was shown. However, the differential reactivi ties of TAP 952 and AXO 49 mAbs with the two axoneme types suggest a d ifference in their glycylation level. In addition, within a single cel l, the spermatozoon, cortical microtubules underlying the flagellar me mbrane, and axonemal microtubules were shown to comprise different tub ulin isoforms, the latter ones only being labelled with one of the ant iglycylated tubulin mAbs, TAP 952. Similarly, the antiacetylated (6-11 B-1) and polyglutamylated (GT335) tubulin mAbs decorated the two types of axonemal microtubules, but not the cortical ones. From these data, a subcellular sorting of posttranslationally modified tubulin isoform s within spermatozoa, on the one hand, and a cellular sorting of glycy lated isoforms inside the whole organism, on the other hand, is demons trated in the flatworm E. caproni. Last, a sequential occurrence of tu bulin posttranslational modifications was observed in the course of sp ermiogenesis. Acetylation appears first, followed shortly by glutamyla tion; glycylation takes place at the extreme end of spermiogenesis and , specifically, in a proximo-distal process. Thus in agreement with, a nd extending other studies [Bre et al., 1996], glycylation appears to close the sequence of posttranslational events occurring in axonemal m icrotubules during spermiogenesis. (C) 1998 Wiley-Liss, Inc.