cr ~ and r ~2 are two heat-and ethanol-inducible g-factors in Escherichia coli. The cr 32 regulon is also induced by unfolded and misfolded proteins in the cytoplasm, and the function of many of the proteins in the cr 32 regulon is to bind to cytoplasmic proteins and assist them in folding or unfolding. To further understand the function of the cr F regulon, we searched for mutants that affected cr E activity. Our results indicate that a signal generated by expression of outer membrane proteins modulates cr E activity. Specifically, r ~ activity is induced by increased expression of OMPs and is reduced by decreased expression of OMPs. In addition, mutations that cause misfolded OMPs induce cr E activity. This signal is generated after the fate of OMPs and periplasmic proteins diverge in the secretory pathway and is not the result of an accumulation of OMP precursors in the cytoplasm. Our results indicate that this effect of OMPs is specific to the r E regnlon, because none of the above mutations affect r 32 activity. We propose that the ~r ~ regnlon is involved in processes that occur in extracytoplasmic compartments and that these two heat-inducible regulons may have distinct but complementary roles of monitoring the state of proteins in the cytoplasm (or 32) and outer membrane ((]rE).[Key Words: (r-Factors; protein export; outer membrane proteins; heat shock; (rE] Received September 13, 1993; revised version accepted October 14, 1993.In bacterial cells the (r-subunit directs RNA polymerase to initiate transcription at promoter sites on the DNA (Burgess et al. 1969). The primary (r-factor in the cell is responsible for transcription of most genes during exponential growth. In addition, alternative (r-factors direct transcription of sets of genes whose products are needed for specific functions, such as sporulation, nitrogen fixation, or flagella synthesis {Gross et al. 1992). Alternative (r-factors are often activated by changes in environmental or cellular conditions that generate morphological and/or molecular cues, signaling the need for the gene products in the regulon under control of a particular (r-factor. Elucidation of these signal-transduction pathways provides insights about global control of gene activity in prokaryotic cells.The activity of two Escherichia coli alternative (r-factors, (r32 and (re ((r24), increases after temperature upshift or exposure to ethanol {Grossman et al. 1984;Erickson et al. 1987;Straus et al. 1987;Erickson and Gross 1989;Wang and Kaguni 1989}. RNA polymerase (E) containing o ~2 (E(r 32) transcribes the heat shock genes with products that consist primarily of chaperones and proteases.

No abstract

The rpoH gene of Escherichia coli encodes CT'^, the 32-kD a-factor responsible for the heat-inducible transcription of the heat shock genes. rpoH is transcribed from at least three promoters. Two of these promoters are recognized by RNA polymerase containing a^", the predominant a-factor. We purified the factor responsible for recognizing the third rpoH promoter [rpoH P3) and identified it as RNA polymerase containing a novel afactor with an apparent M, of 24,000. This new CT, which we call a^, is distinct from the known a factors in molecular weight and promoter specificity, o-^ holoenzyme will not recognize the tr^"-or a^^-controlled promoters we tested, but it does transcribe the htrA gene, which is required for viability at temperatures >42°C. The in vivo role of a^ is not known. The transcripts from the a^-controlled rpoH P3 and htrA promoters are most abundant at very high temperature, suggesting the a^ holoenzyme may transcribe a second set of heatinducible genes that are involved in growth at high temperature or in thermotolerance.

No abstract

The product of the Escherichia coli rpoH (htpR) gene, (F32, is required for heat-inducible transcription of the heat shock genes. Previous studies on the role of &2 in growth at low temperature and in gene expression involved the use of nonsense and missense rpoH mutations and have led to ambiguous or conflicting results. To clarify the role of c32 in cell physiology, we have constructed loss-of-function insertion and deletion mutations in rpoH. Strains lacking r32 are extremely temperature sensitive and grow only at temperatures less than or equal to 20°C. There is no transcription from the heat shock promoters preceding the htpG gene or the groESL and dnaKJ operons; however, several heat shock proteins are produced in the mutants. GroEL protein is present in the rpoH null mutants, but its synthesis is not inducible by a shift to high temperature. The low-level synthesis of GroEL results from transcription initiation at a minor tT70-controlled promoter for the groE operon. DnaK protein synthesis cannot be detected at low temperature, but can be detected after a shift to 42°C. The mechanism of this heat-inducible synthesis is not known. We conclude that cr32 iS required for cell growth at temperatures above 20°C and is required for transcription from the heat shock promoters. Several heat shock proteins are synthesized in the absence of r32, indicating that there are additional mechanisms controlling the synthesis of some heat shock proteins.When cells or organisms are suddenly exposed to high temperature a set of heat shock proteins are transiently induced. The response is apparently universal, having been observed in members of all phylogenetic kingdoms (reviewed in reference 19). In addition, some heat shock proteins have been conserved during evolution. The eucaryotic heat shock proteins Hsp7O and Hsp83 have nearly 50% of their amino acid residues in common with their procaryotic homologs (3, 4). The functions of the heat shock proteins are not well understood. These proteins seem to be involved in a wide variety of cellular processes and are required for prolonged survival at high temperatures (19).In Escherichia coli the heat shock response is rapidly induced. Within 1 min after a shift to high temperature, transcription initiation from the heat shock promoters increases, leading to the elevated synthesis of the heat shock proteins (reviewed in references 24 and 26). The heat shock promoters are recognized in vitro by RNA polymerase containing the 32-kilodalton a subunit (E&2) but not by RNA polymerase containing the primary 70-kilodalton u subunit (E070) (8,10,12). Genetic studies from a number of laboratories support the idea that &32, the product of the rpoH gene, is required for the increased transcription of the heat shock genes after a shift to high temperature. Strains carrying supC(Ts), encoding a temperature-sensitive suppressor tRNA, and the rpoH165 amber mutation fail to induce the synthesis of heat shock proteins and are temperature sensitive for growth (23,26,46 tration of &2 in the cell and t...

the pr0duct 0f the rp0H (htpR) 9ene, 0 ~2, d1rect5 RNA p01ymera5e t0 1n1t1ate tran5cr1pt10n fr0m heat 5h0ck pr0m0ter5 at a11 temperature5. 7ran5cr1pt10n 0f the heat 5h0ck 9ene5 15 1ncrea5ed when ce115 are exp05ed t0 h19h temperature5 6ecau5e 0f 1ncrea5ed tran5cr1pt10n 1n1t1at10n 6y cr32-RNA p01ymera5e. A5 a 5tep t0ward under5tand1n9 the re9u1at10n 0f the heat 5h0ck re5p0n5e we have exam1ned the tran5cr1pt10n 0f the rp0H 9ene. U51n9 51 mapP1n9, pr0m0ter c10n1n9, and 1n v1tr0 tran5cr1pt10n, we have 1dent1f1ed the pr0m0ter5 and the term1nat0r f0r the rp0H tran5cr1pt10n un1t. 7he rp0H tran5cr1pt5 are m0n0c15tr0n1c and 0r191nate fr0m at 1ea5t three pr0m0ter5. N0ne 0f the pr0m0ter5 15 rec09n12ed 6y cr32-RNA p01ymera5e. 7w0 are rec09n12ed 6y (r7°-RNA p01ymera5e and are act1ve at 60th 10w and h19h 9r0wth temperature5. We d0 n0t kn0w what f0rm 0f RNA p01ymera5e rec09n12e5 the th1rd pr0m0ter. 7ran5cr1pt5 fr0m th15 pr0m0ter are a6undant 0n1y at h19h temperature and are pre5ent after 5h1ft t0 the 1etha1 temperature 0f 50°C, even at t1me5 when there are n0 detecta61e tran5cr1pt5 fr0m the 0ther rp0H pr0m0ter5. 7he am0unt 0f rp0H mRNA 1ncrea5e5 f1vef01d 6y 8 m1n after 5h1ft fr0m 30 t0 43.5°C 6ut rp0H mRNA 5ynthe515 1ncrea5e5 6y 1e55 than tw0f01d, 1nd1cat1n9 that there 15 p05t-tran5cr1pt10na1 c0ntr01 0f the 1eve1 0f rp0H mRNA and pre5uma61y 0f cr 32.[Key W0rd5:P05t-tran5cr1pt10na1 re9u1at10n; heat 5h0ck; 519ma-32; rp0H; htpR] When ce115, t155ue5, 0r even wh01e 0r9an15m5 are exp05ed t0 e1evated temperature5 they re5p0nd 6y 1n-crea51n9 the 5ynthe515 0f the heat 5h0ck pr0te1n5. 7h15 re5p0n5e t0 h19h temperature, termed the heat 5h0ck re5p0n5e, 15 51m11ar 1n 0r9an15m5 fr0m every 6101091ca1 k1n9d0m (Ne1dhardt et a1. 1984; L1nd4u15t 1986). 1f ce115 are 5h1fted fr0m a 10w temperature t0 a h19her 0ne w1th1n the1r 9r0wth ran9e, the 1nduct10n 0f the heat 5h0ck pr0te1n5 15 tran51ent. 1f the ce115 are tran5ferred t0 temperature5 a60ve the1r 9r0wth ran9e, the heat 5h0ck pr0te1n5 are expre55ed at h19h 1eve15 f0r a5 10n9 a5 the ce115 5ynthe512e pr0te1n (Ne1dhardt et a1. 1984; L1nd4u15t 1986). 7he funct10n5 0f the heat 5h0ck pr0te1n5 are n0t we11 under5t00d. 1t 15 c1ear that 50me 0f the heat 5h0ck pr0-te1n5 are re4u1red f0r 5urv1va1 at h19h temperature and that expre5510n 0f the heat 5h0ck pr0te1n5 1ncrea5e5 the 1en9th 0f t1me ce115 5urv1ve at temperature5 t00 h19h f0r 9r0wth (Ne1dhardt et a1. 1984; L1nd4u15t 1986). 5t1mu11 0ther than heat can 1nduce the 5ynthe515 0f var10u5 heat 5h0ck pr0te1n5 (Ne1dhardt et a1. 1984; L1nd4u15t 19861. 7he5e 5t1mu11 1nc1ude 5tarvat10n f0r nutr1ent5, v1ra1 1nfect10n, and exp05ure t0 ethan01, 0x1dant5, UV rad1a-t10n, am1n0 ac1d ana1095, 0r DNA-dama91n9 a9ent5. 1t 5eem5 that the heat 5h0ck pr0te1n5 he1p pr0tect ce115 fr0m a var1ety 0f adver5e c0nd1t10n5. 7he heat 5h0ck re5p0n5e 1n E5cher1ch1a c011151nduced rap1d1y. 7he 5ynthe515 0f the 20 0r 50 heat 5h0ck pr0te1n5 15 1ncrea5ed w1th1n 1 m1n after 5h1ft fr0m 30°C t0 a h19her 9r0wth temperature, 5uch a5 43°C (Ne1dhardt et a1. 1984). Max1mum rate5 0f heat 5h0ck pr0te...

In the textbook view, the ratio of X chromosomes to autosome sets, X:A, is the primary signal specifying sexual fate in Drosophila. An alternative idea is that X chromosome number signals sex through the direct actions of several X-encoded signal element (XSE) proteins. In this alternative, the influence of autosome dose on X chromosome counting is largely indirect. Haploids (1X;1A), which possess the male number of X chromosomes but the female X:A of 1.0, and triploid intersexes (XX;AAA), which possess a female dose of two X chromosomes and the ambiguous X:A ratio of 0.67, represent critical tests of these hypotheses. To directly address the effects of ploidy in primary sex determination, we compared the responses of the signal target, the female-specific SxlPe promoter of the switch gene Sex-lethal, in haploid, diploid, and triploid embryos. We found that haploids activate SxlPe because an extra precellular nuclear division elevates total X chromosome numbers and XSE levels beyond those in diploid males. Conversely, triploid embryos cellularize one cycle earlier than diploids, causing premature cessation of SxlPe expression. This prevents XX;AAA embryos from fully engaging the autoregulatory mechanism that maintains subsequent Sxl expression, causing them to develop as sexual mosaics. We conclude that the X:A ratio predicts sexual fate, but does not actively specify it. Instead, the instructive X chromosome signal is more appropriately seen as collective XSE dose in the early embryo. Our findings reiterate that correlations between X:A ratios and cell fates in other organisms need not implicate the value of the ratio as an active signal.

KAP plays an essential role in Kinesin II function, which is required for the axoneme growth and maintenance of the cilia in Drosophila type I sensory neurons. However, the flagellar assembly in Drosophila spermatids does not require Kinesin II and is independent of IFT.