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ABSTRACT: The nuclear pore complex (NPC) mediates all nucleocytoplasmic transport, yet its structure and biogenesis remain poorly understood. In this study, we have functionally characterized interaction partners of the yeast transmembrane nucleoporin Ndc1. Ndc1 forms a distinct complex with the transmembrane proteins Pom152 and Pom34 and two alternative complexes with the soluble nucleoporins Nup53 and Nup59, which in turn bind to Nup170 and Nup157. The transmembrane and soluble Ndc1-binding partners have redundant functions at the NPC, and disruption of both groups of interactions causes defects in Ndc1 targeting and in NPC structure accompanied by significant pore dilation. Using photoconvertible fluorescent protein fusions, we further show that the depletion of Pom34 in cells that lack NUP53 and NUP59 blocks new NPC assembly and leads to the reversible accumulation of newly made nucleoporins in cytoplasmic foci. Therefore, Ndc1 together with its interaction partners are collectively essential for the biosynthesis and structural integrity of yeast NPCs.
The Journal of Cell Biology 06/2009; 185(3):475-91. · 10.26 Impact Factor
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ABSTRACT: We have established that two homologous nucleoporins, Nup170p and Nup157p, play an essential role in the formation of nuclear pore complexes (NPCs) in Saccharomyces cerevisiae. By regulating their synthesis, we showed that the loss of these nucleoporins triggers a decrease in NPCs caused by a halt in new NPC assembly. Preexisting NPCs are ultimately lost by dilution as cells grow, causing the inhibition of nuclear transport and the loss of viability. Significantly, the loss of Nup170p/Nup157p had distinct effects on the assembly of different architectural components of the NPC. Nucleoporins (nups) positioned on the cytoplasmic face of the NPC rapidly accumulated in cytoplasmic foci. These nup complexes could be recruited into new NPCs after reinitiation of Nup170p synthesis, and may represent a physiological intermediate. Loss of Nup170p/Nup157p also caused core and nucleoplasmically positioned nups to accumulate in NPC-like structures adjacent to the inner nuclear membrane, which suggests that these nucleoporins are required for formation of the pore membrane and the incorporation of cytoplasmic nups into forming NPCs.
The Journal of Cell Biology 06/2009; 185(3):459-73. · 10.26 Impact Factor
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ABSTRACT: Structural aspects of lipoarabinomannans (LAM) from Mycobacterium tuberculosis and Mycobacterium smegmatis were investigated by using mild acid hydrolysis in combination with Fourier-transform ion cyclotron resonance (FT-ICR), and quadrupole ion trap mass spectrometry. Exact mass measurements with less than 2.5 ppm mass error confirmed the presence of a series of arabinose oligomers (Ara(n); n = 2-7) as the major components observed following mild acid hydrolysis of both M. tuberculosis and M. smegmatis LAM. However, the mass spectrum of the resulting LAM extract also revealed a highly-abundant distribution of ions that exact mass measurements identified as mannose-linked arabinose species, Ara(n)Man(m) + Na+ (n = 1-6; m = 1-3). The observed mannose caps were linked to arabinose species as mono-, di-, and trimannose units, and the ratio of the mono-, di-, and trimannose caps was determined to be 1.00:9.00:1.15, respectively, different from previous reports. Analysis of the linkage of lithiated arabinose trimer standards was accomplished with MS3 experiments and the information generated was used to identify linkages of arabinose trimers generated by mild acid hydrolysis of M. tuberculosis and M. smegmatis LAM. The MS3 spectra confirmed the linkage of arabinose trimers from M. smegmatis and M. tuberculosis LAM as predominantly alpha(1 --> 5), alpha(1 --> 5).
Journal of the American Society for Mass Spectrometry 07/2005; 16(7):1109-16. · 4.00 Impact Factor
