Glutamylated and glycylated tubulin isoforms in the aberrant sperm axoneme of the gall‐midge fly, Asphondylia ruebsaameni

Abstract

The axonemal organization expressed in the sperm flagella of the cecidomyiid dipteran Asphondylia ruebsaameni is unconventional, being characterized by the presence of an exceedingly high number of microtubular doublets and by the absence of both the inner dynein arms and the central pair/radial spoke complex. Consequently, its motility, both in vivo and in vitro, is also peculiar. Using monoclonal antibodies directed against posttranslational modifications, we have analyzed the presence and distribution of glutamylated and glycylated tubulin isoforms in this aberrant axonemal structure, and compared them with those of a reference insect species (Apis mellifera), endowed with a conventional axoneme. Our results have shown that the unorthodox structure and motility of the Asphondylia axoneme are concomitant with: (1). a very low glutamylation extent in the alpha-tubulin subunit, (2). a high level of glutamylation in the beta-subunit, (3). an extremely low total extent of glycylation, with regard to both monoglycylated and polyglycylated sites, either in alpha- or in beta-tubulin, (4). the presence of a strong labeling of glutamylated tubulin isoforms at the proximal end of the axoneme, and (5). a uniform distribution of glutamylated as well as glycylated isoforms along the rest of the axoneme. Thus, our data indicate that tubulin molecular heterogeneity is much lower in the Asphondylia axoneme than in the conventional 9+2 axoneme with regard to both isoform content and isoform distribution along the axoneme.

The proximal region of the β-tubulin C-terminal tail is sufficient for axoneme assembly

“…We conclude that in Drosophila only part of the total axonemal tubulins are modified. This is consistent with observations that only a fraction of the total axonemal tubulin is modified in many other organisms [Kann et al, 1995;Mary et al, 1996Mary et al, , 1997Multigner et al, 1996;Weber et al, 1997;Schneider et al, 1998;Mencarelli et al, 2000Mencarelli et al, , 2004. The stability tests and gel analyses presented above suggest that b2D7 has biochemical properties similar to full-length b2-tubulin, but has slightly decreased capacity to associate with a-tubulin.…”

Tubulin glycylation and glutamylation deficiencies in unconventional insect axonemes

“…5A), reminiscent of that found in Asphondylia ruebsaameni [Mencarelli et al, 2004] (see Discussion). It consists of 170-200 microtubular doublets arranged in different laminae, each one extending with a grossly helical disposition from about the centre of the flagellum to the periphery and the plasma membrane.…”

“…We have recently described the tubulin isoform content of the aberrant axoneme present in the sperm flagella of the dipteran cecidomyiid Asphondylia ruebsaameni [Mencarelli et al, 2004]. In this species, the giant sperm displays an unorthodox axonemal pattern, which consists of a huge number of microtubular doublets and is devoid of both the central pair/radial spoke complex and the inner dynein arms.…”

“…We have recently analyzed the flagellar tubulin isoform content in a dipteran insect, the cecidomyiid Asphondylia, which is characterized by an aberrant axonemal structure, and have shown that it is characterized by a reduced tubulin molecular heterogeneity [Mencarelli et al, 2004]. Here, we extend our study to the analysis of glycylated and glutamylated tubulin isoforms in the sperm flagellum of other insect species possessing a modified axonemal organization; three of them exhibit axonemes devoid of the central microtubular complex, while the fourth one shows a completely aberrant axonemal structure.…”

Axoneme‐dependent tubulin modifications in singlet microtubules of the Drosophila sperm tail

“…They found that the majority of polymodified tubulins were in the axonemal microtubule fraction, not in the accessory microtubules, suggesting greater tubulin heterogeneity in axonemal microtubules than in the accessory tubules. Mencarelli and colleagues also found that compared to the standard axonemes of Drosophila and honey bee sperm, there was substantially less C-terminal modified tubulin in insect species with sperm axonemes lacking central pairs [Mencarelli et al, 2004[Mencarelli et al, , 2005. These included 9 1 9 1 0 axonemes in several insects that have sperm with standard architecture of nine doublets plus nine accessory microtubules but no central pairs (a mosquitoe, Anopheles stephensi; a mayfly, Ecdyonurus venosus; and a hawthorn fly, Bibio marci), and the remarkable variant axonemes of the gall-midge fly Asphondylia ruebsaameni and the leafminer Monarthropalpus flavus, composed only of huge complex arrays of doublet microtubules.…”