Equivalent Mutations in the Eight Subunits of the Chaperonin CCT Produce Dramatically Different Cellular and Gene Expression Phenotypes
JOURNAL OF MOLECULAR BIOLOGY
Authors: Amit, Maya; Weisberg, Sarah J.; Nadler-Holly, Michal; McCormack, Elizabeth A.; Feldmesser, Ester; Kaganovich, Daniel; Willison, Keith R.; Horovitz, Amnon
Abstract
The eukaryotic cytoplasmic chaperonin-containing TCP-1 (CCT) is a complex formed by two back-to-back stacked hetero-octameric rings that assists the folding of actins, tubulins, and other proteins in an ATP-dependent manner. Here, we tested the significance of the hetero-oligomeric nature of CCT in its function by introducing, in each of the eight subunits in turn, an identical mutation at a position that is conserved in all the subunits and is involved in ATP hydrolysis, in order to establish the extent of 'individuality' of the various subunits. Our results show that these identical mutations lead to dramatically different phenotypes. For example, Saccharomyces cerevisiae yeast cells with the mutation in subunit CCT2 display heat sensitivity and cold sensitivity for growth, have an excess of actin patches, and are the only strain here generated that is pseudo-diploid. By contrast, cells with the mutation in subunit CCT7 are the only ones to accumulate juxtanuclear protein aggregates that may reflect an impaired stress response in this strain. System-level analysis of the strains using RNA microarrays reveals connections between CCT and several cellular networks, including ribosome biogenesis and TOR2, that help to explain the phenotypic variability observed. (C) 2010 Elsevier Ltd. All rights reserved.
Sequencing analysis of a 40 center dot 2 kb fragment of yeast chromosome X reveals 19 open reading frames including URA2 (5' end), TRK1, PBS2, SPT10, GCD14, RPE1, PHO86, NCA3, ASF1, CCT7, GZF3, two tRNA genes, three remnant delta elements and a Ty4 transposon
YEAST
Authors: Cziepluch, C; Kordes, E; Pujol, A; Jauniaux, JC
Abstract
The complete sequence of a 40 247 bp DNA segment located on the left arm of chromosome X of Saccharomyces cerevisiae has been determined and analysed. The sequence encodes the 5' coding region of the URA2 gene and 18 open reading frames of at least 100 amino acids. Ten of these correspond to known genes, whereas eight correspond to new genes. In addition, the sequence contains a tRNA-Ala gene, a tRNA-Asp gene, a Ty4 transposable element and three delta elements. The sequence has been deposited in the EMBL databank under Accession Numbers: Z49405, Z49404, Z49403, Z49402, Z49401, Z49400, Z49399, Z49398, Z49397, Z49396, Z49394, Z49392, Z49391, Z49390, Z49389, Z49387, Z49386, Z49385.
COA
Certificate of Analysis
