Sex Plant Reprod (1995) 8:326-332 9 Springer-Verlag 1995 R. Kulikauskas 9 A. Hou 9 J. Muschietti 9 S. McCormick Comparisons of diverse plant species reveal that only grasses show drastically reduced levels of ubiquitin monomer in mature pollen Received: 5 July 1995 / Accepted: 20 July 1995 Abstract Ubiquitin is a ubiquitous protein involved in targeting proteins for degradation. Maize pollen was pre- viously reported (Callis and Bedinger 1994) to show ex- tremely low levels of ubiquitin monomer, and develop- mental significance was attributed to this surprising fea- ture of maize pollen. However, we had previously shown (Muschietti et al. 1994) that tomato pollen had high lev- els of ubiquitin monomer. Here we show that pollen from most plant families has high levels of ubiquitin monomer. Most grasses tested show reduced levels of ubiquitin monomer, but some maize inbred lines have higher levels of ubiquitin monomer than other inbreds. There was no correlation between the level of ubiquitin monomer and either the monocotyledonous or tri-cellular condition of grass pollen or the dehydrated condition of mature pollen. Since many aspects of pollen develop- ment (i.e., wall formation, microspore mitosis, synthesis and storage of mRNAs and proteins, carbohydrates and lipids, dehydration at maturity) are stereotypical among all plant families, the reduced level of ubiquitin mono- mer in pollen of many grasses cannot be crucial for any feature of normal pollen development. Key words Ubiquitin 9 Tomato - Maize - Heat shock proteins 9 Protein degradation Introduction Pollen development is strictly oriented toward its final function, fertilization of the female gametophyte (re- viewed in Mascarenhas 1993; McCormick 1993). Pollen has specific and tight control of its gene expression, pro- ducing the required proteins at the right time (reviewed in Mascarenhas 1989). For example, pollen genes acti- R. Kulikauskas 9 A. Hou 9 J. Muschietti 9 S. McCormick ( ~ ) Plant Gene Expression Center, U.S. Department of Agriculture - Agricultural Research Service and University of California at Berkeley, 800 Buchanan Street, Albany, CA 94710, USA; Fax no.: (510) 559-5678 vated before microspore mitosis are thought to be re- quired for "early" events in pollen development, such as cytoskeletal organization and pollen cell wall formation (Evans et al. 1992), while "late genes" that are first ex- pressed after microspore mitosis and peak in mature pol- len are thought to be involved in pollen maturation or pollen tube growth (Muschietti et al. 1994). While it is reasonable to assume that pollen proteins are degraded after their function has been accomplished, there are no conclusive reports as to whether developmentally con- trolled protein turnover is important in pollen. Ubiquitin is a highly conserved 76-amino-acid protein found in all eukaryotic cells and is involved in the most well-characterized protein degradation pathway. Ubiqui- tin is found free or covalently bound to other cytoplas- mic, nuclear or integral membrane proteins. Once multi- ple ubiquitin moieties are bound, the target protein is rapidly degraded by a protease complex, with the subse- quent liberation and reuse of the free ubiquitin (reviewed in Hershko and Ciechanover 1992). Ubiquitin and many ubiquitin genes and ubiquitin pathway enzymes or the genes encoding them have been isolated from plants (re- viewed in Vierstra 1993). The patterns of ubiquitinated proteins change during processes such as floral develop- ment (Courtney et al. 1994), zygotic embryogenesis (A1- moguera et al. 1995), aging and greening of leaves (Ve- ierskov et al. 1992) and seed formation (Ferreira et al. 1995), suggesting that ubiquitin-mediated protein degra- dation plays a role during all of these processes. It was recently shown that free and protein-bound ubiquitin levels - but not ubiquitin mRNA - are dramat- ically reduced during maize pollen development (Callis and Bedinger 1994). The timing of this reduction corre- lated with the initiation of microspore mitosis and pol- len maturation. Because of their finding with maize pol- len, Callis and Bedinger suggested that a tight regula- tion of protein turnover might not be required in mature pollen because it has a short life span. They also pro- posed that ubiquitination of proteins could be detrimen- tal during pollen dehydration and that ubiquitin protein levels might be down-regulated in order to prevent erro-