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TPPP/p25 promotes tubulin assemblies and blocks mitotic spindle formation

2003; National Academy of Sciences; Volume: 100; Issue: 24 Linguagem: Inglês

10.1073/pnas.2436331100

1091-6490

László Tirián, Emma Hlavanda, Judit Oláh, István Horváth, Ferenc Orosz, Bálint Sámuel Szabó, Joseph A. Kovacs, János Szabad, Judit Ovádi,

Ubiquitin and proteasome pathways

Recently, we isolated from bovine brain a protein, TPPP/p25 and identified as p25, a brain-specific protein that induced aberrant tubulin assemblies. The primary sequence of this protein differs from that of other proteins identified so far; however, it shows high homology with p25-like hypothetical proteins sought via blast . Here, we characterized the binding of TPPP/p25 to tubulin by means of surface plasmon resonance; the kinetic parameters are as follows: k on , 2.4 × 10 4 M –1 ·s –1 ; k off , 5.4 × 10 –3 s –1 ; and K d , 2.3 × 10 –7 M. This protein at substoichometric concentration promotes the polymerization of tubulin into double-walled tubules and polymorphic aggregates or bundles paclitaxel-stabilized microtubules as judged by quantitative data of electron and atomic force microscopies. Injection of bovine TPPP/p25 into cleavage Drosophila embryos expressing tubulin–GFP fusion protein reveals that TPPP/p25 inhibits mitotic spindle assembly and nuclear envelope breakdown without affecting other cellular events like centrosome replication and separation, microtubule nucleation by the centrosomes, and nuclear growth. GTP counteracts TPPP/p25 both in vitro and in vivo .