Interactions among Drosophila Nuclear Envelope Proteins Lamin, Otefin, and YA

ABSTRACT

The nuclear envelope plays many roles, including organizing nuclear structure and regulating nuclear events. Molecular associations of nuclear envelope proteins may contribute to the implementation of these functions. Lamin, otefin, and YA are the three Drosophilanuclear envelope proteins known in early embryos. We used the yeast two-hybrid system to explore the interactions between pairs of these proteins. The ubiquitous major lamina protein, lamin Dm, interacts with both otefin, a peripheral protein of the inner nuclear membrane, and YA, an essential, developmentally regulated protein of the nuclear lamina. In agreement with this interaction, lamin and otefin can be coimmunoprecipitated from the vesicle fraction ofDrosophila embryos and colocalize in nuclear envelopes ofDrosophila larval salivary gland nuclei. The two-hybrid system was further used to map the domains of interaction among lamin, otefin, and YA. Lamin’s rod domain interacts with the complete otefin protein, with otefin’s hydrophilic NH₂-terminal domain, and with two different fragments derived from this domain. Analogous probing of the interaction between lamin and YA showed that the lamin rod and tail plus part of its head domain are needed for interaction with full-length YA in the two-hybrid system. YA’s COOH-terminal region is necessary and sufficient for interaction with lamin. Our results suggest that interactions with lamin might mediate or stabilize the localization of otefin and YA in the nuclear lamina. They also suggest that the need for both otefin and lamin in mediating association of vesicles with chromatin might reflect the function of a protein complex that includes these two proteins.

ACKNOWLEDGMENTS

Michal Goldberg and Huihua Lu contributed equally to this work.

We thank T. Fox, J. Hirschberg, M. L. Goldberg, and D. Starr for helpful comments on this paper; T. Fox and his lab for providing yeast strains, media, and advice to H.L. and M.F.W.; and D. Scott for assistance with the cloning of YA506-696. M.F.W. also thanks B. Wakimoto and her lab for their hospitality and advice, R. Wright and L. Hartwell for advice and media, and S. Turner for initial PCRs during the construction of and initial experiments with the full-length two-hybrid fusions.

This work was supported by grants from NIH (GM44659 to M.F.W. and GM33132 to P.A.F.) and the Israel Academy of Sciences (Y.G.) and by long-term fellowships from the Human Frontiers Science Program Organization and EMBO (N.S.).