The first gene in the Escherichia coli secA operon, gene X, encodes a nonessential secretory protein

We describe an Escherichia coli genetic screen that yields mutations affecting two different cellular processes: disulfide bond formation and membrane protein assembly. The mutants defective in disulfide bond formation include additional classes of dsbA and dsbB mutations. The membrane protein assembly defective mutants contain a mutation in the secA operon and three mutations in the ffs gene, which encodes 4.5S RNA. These latter mutations are the only ones to be isolated in a gene encoding a component of the bacterial signal recognition particle by screening in vivo for defects in membrane protein insertion. A sensitive method for examining membrane protein localization shows that the ffs and secA locus mutations affect membrane assembly of the polytopic membrane protein, MalF. The ffs mutations also affect the membrane insertion of the FtsQ and the AcrB proteins. Although both the ffs and the secA locus mutations interfere with membrane protein assembly, only the latter also reduces export of a protein containing a cleavable signal sequence.

Section: Resultsmentioning

confidence: 78%

A mutant hunt for defects in membrane protein assembly yields mutations affecting the bacterial signal recognition particle and Sec machinery

Tian

Boyd

Beckwith

2000

“…This regulation is at the translation level, in which secM (secretion monitor) located upstream of secA in the secM-secA-mutT polycistronic messenger RNA (8) plays a crucial role (9). The product of secM is a peculiar secretory protein (10,11), which undergoes translation elongation-arrest at the Pro-166 position of the 170-codon-long secM ORF (12). Thus, ribosome-tethered SecM 1-166 -tRNA accumulates transiently in normal cells and more stably in secretion-defective cells (11,12).…”

mentioning

confidence: 99%

Translation arrest of SecM is essential for the basal and regulated expression of SecA

Murakami

Nakatogawa

Ito

2004

The SecM protein of Escherichia coli contains an arrest sequence (F 150 XXXXWIXXXXGIRAGP 166 ), which interacts with the ribosomal exit tunnel to halt translation elongation beyond Pro-166. This inhibition is reversed by active export of the nascent SecM chain. Here, we studied the physiological roles of SecM. Arrest-alleviating mutations in the arrest sequence reduced the expression of secA, a downstream gene on the same mRNA. Among such mutations, the arrest-abolishing P166A substitution mutation on the chromosomal secM gene proved lethal unless the mutant cells are complemented with excess SecA. Whereas secretion defect due either to azide addition, a secY mutation, or low temperature leads to up-regulated SecA biosynthesis, this regulation was lost by a secM mutation, which synergistically retarded growth of cells with lowered secretion activity. Finally, an arrest-alleviating rRNA mutation affecting the constricted part of the exit tunnel lowered the basal level of SecA as well as its secretion defect-induced upregulation. Thus, the arrest sequence of SecM has at least two roles in SecA translation. First, the transient elongation arrest in normal cells is required for the synthesis of SecA at levels sufficient to support cell growth. Second, the prolonged SecM elongation arrest under conditions of unfavorable protein secretion is required for the enhanced expression of SecA to cope with such conditions.

“…E. coli SecM is encoded by the 170-codon open reading frame upstream of secA, encoding the protein export-driving ATPase, in the same transcription unit (8)(9)(10). It is designed as a periplasmic protein but contains an arrest sequence, F 150 XXXXWIXXXXGIRAGP 166 , which was suggested to interact with the exit tunnel of the ribosome (6).…”

mentioning

confidence: 99%

Recruitment of a species-specific translational arrest module to monitor different cellular processes

Chiba

Kanamori

Ueda

et al. 2011

Nascent chain-mediated translation arrest serves as a mechanism of gene regulation. A class of regulatory nascent polypeptides undergoes elongation arrest in manners controlled by the dynamic behavior of the growing chain; Escherichia coli SecM monitors the Sec protein export pathway and Bacillus subtilis MifM monitors the YidC membrane protein integration/folding pathway. We show that MifM and SecM interact with the ribosome in a speciesspecific manner to stall only the ribosome from the homologous species. Despite this specificity, MifM is not exclusively designed to monitor membrane protein integration because it can be converted into a secretion monitor by replacing the N-terminal transmembrane sequence with a secretion signal sequence. These results show that a regulatory nascent chain is composed of two modular elements, one devoted to elongation arrest and another devoted to subcellular targeting, and they imply that physical pulling force generated by the latter triggers release of the arrest executed by the former. The combinatorial nature may assure common occurrence of nascent chain-mediated regulation.stalling sequence | SpoIIIJ | SecA | exit tunnel