FIG. 5.

Hmt1 influences nucleocytoplasmic transport of C. albicans Npl3. (A) NPL3 was GFP tagged by homologous recombination in homozygous C. albicans strains with (+; AMC46 and AMC47) or without (−; AMC48 and AMC49) Hmt1. PCR products used for recombination contained either wild-type NPL3 sequences (WT) or introduced a T-to-G mutation in codon 340, encoding an S-to-A (SA) mutation in the C terminus of Npl3. Equal amounts of total protein from mid-log-phase GFP-tagged and parental cells were immunoprecipitated with anti-GFP and PrG-Sepharose. Equal amounts of each precipitate were analyzed by immunoblotting with anti-GFP (from 0.3 mg total protein) and antidimethylarginine antiserum (from 0.6 mg total protein). Parental strains lacking GFP (−; BWP17 [+Hmt1] and AMC36 [−Hmt1]) and an HMT1/HMT1 strain expressing GFP alone (G; MLR62) were used as controls. (B) HMT1/HMT1 (HMT1+; AMC46, AMC47) and hmt1Δ/hmt1Δ (HMT1−; AMC48 and AMC49) strains were grown to mid-log phase and stained with DAPI. Cells were visualized by Nomarski and fluorescence microscopy using GFP and DAPI filters. For each filter, exposure times were equivalent for all strains.