Localization of proteins that are coordinately expressed with Cln2 during the cell cycle

Abstract

The localization of proteins can give important clues about their function and help sort data from large-scale proteomic screens. Forty-five proteins were tagged with the GFP variant YFP. These proteins were chosen because they are encoded by genes that display strong cell cycle-dependent expression that peaks in G 1 . Most of these proteins localize to either the nucleus or to sites of cell growth. We are able to assign new cellular component GO terms to ASF2, TOS4, RTT109, YBR070C, YKR090W, YOL007C, YOL019W and YPR174C. We also have localization data for 21 other proteins. Noteworthy localizations were found for Rfa1p, a member of the DNA replication A complex, and Pri2p and Pol12p, subunits of the α-DNA polymerase : primase complex. In addition to its nuclear localization, Rfa1p assembled into cytoplasmic foci adjacent to the nucleus in cells during the G 1 -S phase transition of the cell cycle. Pri2 and Pol12 took on a beaded appearance at the G 1 -S transition and later in the cell cycle were enriched in the nuclear envelope. A new spindle pole body/nuclear envelope component encoded by YPR174 was identified. The cell cycle-dependent abundance of Tos4p mirrored Yox1p and these two proteins were the only proteins that were found exclusively at the G 1 -S phase of the cell cycle. A complete list of localizations, along with images, can be found at our website (http://www.yeastrc.org/cln2/).

TheSaccharomyces cerevisiaeSpindle Pole Body (SPB) Component Nbp1p Is Required for SPB Membrane Insertion and Interacts with the Integral Membrane Proteins Ndc1p and Mps2p

“…Previous studies have shown that Nbp1p is present as one or two spots at the nuclear periphery (Shimizu et al, 2000) and colocalizes with the SPB component Spc29p (Sundin et al, 2004). This localization is consistent with an association of Nbp1p either with SPBs or with kinetochores, which in yeast are clustered close to SPBs.…”

“…In addition, the protein encoded by S. cerevisiae YPR174C displays significant homology to the C-terminal half (residues 188-304) of Nbp1p (35% identical, 52% similar). Like Nbp1p, the protein encoded by YPR174C also contains a coiled-coil domain and has been localized to the SPB and nuclear periphery by fluorescence microscopy (Huh et al, 2003;Sundin et al, 2004). Both promoters of NBP1 and YPR174C contain a MluI "cell cycle box" and in both cases the mRNA expression peaks during G1 (Spellman et al, 1998;Shimizu et al, 2000), consistent with playing a role during SPB duplication.…”

The Organization of the Core Proteins of the Yeast Spindle Pole Body

“…The compositions of synthetic dextrose minimal medium (SD), SD-complete (SDC), and yeast peptone dextrose-rich broth medium (YPD) are described elsewhere (Sundin et al, 2004).…”

“…Microcolonies were scraped off the plate and suspended in 30 l of SDC medium. A 3-4-l aliquot was mounted on a pad of 0.9% SeaKem LE agarose (FMC BioProducts, Rockland, ME) in SDC as described (Sundin et al, 2004).…”

Retroviruses and yeast retrotransposons use overlapping sets of host genes

“…1I). Rtt109 is nuclear and has an RNA expression pattern similar to that of Cln2 (Sundin et al 2004), but its function is not known. The mechanism through which DNA maintenance functions affect Ty1 cDNA is not completely understood but, given the common proteins and similar phenotypes, is likely to operate in the case of Ty3 as well.…”