Hemocyte responses to implanted tissues inDrosophila melanogaster larvae

Summary At 26° C temperature-sensitivetu(1) Sz ^ts larvae ofDrosophila melanogaster develop melanotic tumors consisting of aberrant caudal adipose tissue encapsulated by precociously differentiated hemocytes (lamellocytes). Whentu-Sz ^ts larvae are grown at 18° C, lamellocytes are present but the caudal fat body surfaces remain normal and melanotic tumors do not develop (Rizki and Rizki, preceding paper). In this paper we demonstrate that the lamellocytes intu-Sz ^ts larvae at 18° C encapsulate implants of mechanically-damaged fat bodies and adipose cells devoid of basement membrane, while leaving host fat bodies or implanted fat bodies with intact basement membrane unencapsulated. Therefore, low temperature blocks melanotic tumor formation by normalizing the surfaces of the prospective tumor-forming sites intu-Sz ^ts.The discriminatory ability oftu-Sz ^ts lamellocytes was examined by challenging them with undamaged heterospecific tissues. Tissues from sibling species ofD. melanogaster were not encapsulated whereas tissues fromDrosophila species outside theD. melanogaster species subgroup were. Ultrastructural examination of encapsulated heterospecific tissues showed intact basement membrane, so we propose that distinction between self and not self by lamellocytes depends upon the molecular architecture of the basement membrane. In similar series of experiments usingD. virilis donor tissues inOre-R wild type larval hosts, fat bodies remained unencapsulated and imaginal disks metamorphosed. These studies suggest that continued presence of lamellocytes in the larval host is a prerequisite for encapsulation.     

Hemocyte reactions and early cellular changes during melanotic tumor formation in Drosophila melanogaster

In Drosophila melanogaster tu bw larvae melanotic tumors form as a result of a cell-mediated immune response involving the encapsulation and melanization by hemocytes of portions of the caudal adipose tissue. The tissue-specific encapsulation response is not due to the disintegration of the basement membrane surrounding the adipose tissue as is reported to be the case in other melanotic mutants. Prior to encapsulation large numbers of hemocytes appear in the circulation and begin to differentiate into flattened cells termed lamellocytes. This transformation occurs at a time when changes are noted within the adipose cells. The localized accumulation of blood cells near intact basement membrane suggests that abnormally developing adipose cells acquire altered molecular surfaces or release substances to which the hemocytes respond. The initial reaction of the hemocytes with the adipose tissue is cell lysis, and this is rapidly followed by the aggregation of numerous additional blood cells which eventually cohere to one another to form a multilayered capsule. What little evidence there is of disintegration of the basement membrane occurs only after hemocytes have lysed at the surface, and other blood cells begin to invade the adipose tissue. Melanization occurs first in the intercellular spaces along the plasma membranes of the lysed cells, and progresses from the innermost layers toward the periphery of the encapsulating cells. Since the changes observed in the hemocytes and adipose cells are precocious, occurring to a lesser degree later in normal development, the initial effect of the genetic mutation in tu bw larvae may be an endocrine dysfunction which causes an asynchronous and abnormal development of the caudal adipose tissue and/or the hematopoietic organs.     

Isolation and characterization of temperature-sensitivelethal(2)giant larva alleles

Summary In this paper we present an analysis of the behavior ofl(2)gl ^tsimaginal wing discs during culture in adult hosts. Thel(2)gl ^tslarvae reared at 29° C contain two types of wing discs, those that are morphologically normal and those that are abnormal. When discs of both types are cultured in adult hosts at 29° C, the restrictive temperature, they give rise to transplantable neoplastic tissue. However, when the 29° C reared discs are cultured at 15° C, the permissive temperature, the morphologically normal discs maintain their morphology, but the morphologically abnormal discs give rise to neoplasms. Thel(2)gl ^tslarvae reared at 15° C contain only morphologically normal discs. When these discs are cultured in adult hosts at 29° C they give rise to neoplasms, however if the discs are cultured at 15° C they maintain their normal morphology. These results demonstrate: (1) that all wing imaginal discs obtained from 29° C rearedl(2)gl ^tslarvae are competent to undergo neoplastic development, (2) the morphologically abnormal discs obtained from the 29° C rearedl(2)gl ^tslarvae are committed to neoplastic development, (3) the neoplastic development of the morphologically normal discs is temperature dependent, (4) once the neoplastic development of thel(2)gl ^tsdiscs has been initiated the process is not readily reversible. In addition, the ability ofl(2)gl ^tswing discs to perform epimorphic regulation was tested by amputating morphologically normal permissively rearedl(2)gl ^tswing discs and culturing both fragiments at the permissive temperature. Fragments of control wild-type discs maintained their morphology during culture at the permissive temperature. However, both fragments of txel(2)gl ^tsdiscs became neoplastic. This result is discussed with respect to a possible role for thel(2)gl ⁺function in epimorphic regulation and with respect to the phenomena of tumor promotion in vertebrates.     

Isolation and characterization of immunity temperature sensitive mutants of Enterobacter cloacae harbouring the cloacinogenic factor DF13

Summary Enterobacter cloacae cells, harbouring the cloacinogenic factor DF13 (Clo DF13) are immune to the cloacin they produce. We describe the isolation of eleven Enterobacter cloacae (Clo DF13) mutants, which are immune at 30°C, but lose their immunity at 42°C. The temperature sensitive immunity (Imm^ts) of these mutants appeared not to be transferable together with the Clo DF13 factor to non-cloacinogenic acceptor strains. Apparently host mutations are involved in the Imm^ts phenotype. Two different groups of Imm^ts mutants could be identified. Imm^tsC6 and Imm^tsC8, representatives of each group, have been compared with the parent strain. Imm^tsC6 as well as Imm^tsC8 is sensitive to crude cloacin at 42°C. Imm^ts mutants appeared to be also sensitive to cell components other than cloacin, indicating that the Imm^ts mutations may result in pleiotropic changes of cell properties.The Imm^tsC6 mutant is sensitive to deoxycholate and osmotic shock at 42°C. Spheroplasts of Imm^tsC6 cells incubated at 42°C are sensitive to DOC at 42°C and 30°C. The pleiotrophic changes of the Imm^tsC6 mutant may be attributed to a defect in the cell membrane.The Imm^tsC8, incubated at 42°C, is sensitive to deoxycholate, osmotic shock, ethylene-diaminetetraacetic acid, dyes, drugs and UV. Furthermore they form filaments. Imm^tsC8 spheroplasts are as sensitive to deoxycholate as the parent strain at 42°C. The pleiotropic changes in the phenotype of Imm^tsC8 are considered to be the result of a defect in the outer layers of the cell envelope, most likely the lipopolysaccharide layer.The possible relationship between the observed structural defects in the cell envelope of Imm^ts mutants and the phenomenon of immunity have been discussed.     

Lethal(1)aberrant immune response mutations leading to melanotic tumor formation in Drosophila melanogaster

Using P element-mediated mutagenesis we have isolated 20 X-linked lethal mutations, representing at least 14 complementation groups, which exhibit melanotic tumor phenotypes. We present the systematic analysis of this interesting group of lethal mutations that were selected for their visible melanotic or immune response. The lethal and melanotic tumor phenotypes of each lethal(1) aberrant immune response (air) mutation are pleiotropic effects of single genetic lesions. Lethality occurs throughout the larval and early pupal periods of development and larval development is extended in some air mutants. The air mutant lethal syndromes include abnormalities associated with the brain, haematopoietic organs, gut, salivary glands, ring glands, and imaginal discs. Additional characterization of the melanotic tumor mutations Tum^l and tu(1)Sz^ts have indicated that the melanotic tumor phenotype is similar to that observed in the air mutants. These studies have led to the proposal that two distinct classes of melanotic tumor mutations exist. Class 1 includes mutants in which melanotic tumors result from “autoimmune responses” or the response of an apparently normal immune system to the presence of abnormal target tissues. The Class 2 mutants display obvious defects in the haematopoietic organs or haemocytes, manifested as overgrowth, and the resulting aberrant immune system behavior may contribute to melanotic tumor formation.     

Leptopilina heterotoma and L. boulardi: strategies to avoid cellular defense responses of Drosophila melanogaster

Eggs of three strains of the cynipid parasitoid Leptopilina heterotoma and a Tunisian strain (G317) of L. boulardi are not encapsulated by hemocytes of Drosophila melanogaster hosts, but the eggs of a Congolese strain (L104) of L. boulardi are encapsulated. To determine the reason for the difference in host response against the parasitoid eggs, lamellocytes (hemocytes that encapsulate foreign objects and form capsules around endogenous tissues in melanotic tumor mutants) were examined in host larvae parasitized by the five Leptopilina strains. Parasitization by the three L. heterotoma strains affected the morphology of host lamellocytes and suppressed endogenous melanotic capsule formation in melanotic tumor hosts. L104 did not alter the morphology of host lamellocytes nor block tumor formation in melanotic tumor mutant hosts. The morphology of some lamellocytes was affected by G317 parasitization but host lamellocytes were still capable of forming melanotic tumors and encapsulating dead supernumerary parasitoid larvae. Therefore, the eggs of strains affecting lamellocyte morphology are protected from encapsulation by the host's blood cells. L. heterotoma eggs float freely in the host hemocoel but L. boulardi eggs are attached to host tissue surfaces. Lamellocytes cannot infiltrate the attachment site so the capsule around the L104 egg remains incomplete. The wasp larva uses this gap in the capsule as an escape hatch for emergence.     

Heat-sensitive DNA-binding activity of thecI product of bacteriophage lambda

Summary The binding of genecI product to DNA was studied at temperatures from 0°C to 46° C. Binding activity of the products ofcIts mutants was higher at 22° C than at 0° C, 26° C or 30° C. BothcI⁺ andcIts products lost DNA-binding activity at 46° C, but after subsequent cooling to 22° C, they regained 50–100% of their activity.     

Developmental genetics of a temperature-sensitive RNA polymerase II mutation in Drosophila melanogaster

Summary Purified RNA polymerase II (RNA nucleotidyl-transferase; EC 2.7.7.6) extracted from flies possessing lesions in the Ultrabithorax-like (Ubl) locus of Drosophila melanogaster has altered activity in vitro (Greenleaf et al. 1979, 1980; Coulter and Greenleaf 1982). This strongly suggests that the Ubl locus encodes a subunit of RNA polymerase II. Ethyl methanesulfonate was used to induce a temperature-sensitive mutation in this locus. Flies either homozygous or hemizygous for this new X–linked mutation (Ubl ^ts) display viability comparable to that of wild-type flies at 22° C but are lethal at 29° C. The temperature-sensitive period for Ubl ^ts flies is between gastrulation (6 h, 29° C) and pupation (9–10 days, 22° C). Zygotes shifted from 22° C to 29° C die at either the late embryonic or first larval instar stage while temperature shifts of second and third instar larvae result in the lethal phase occurring at the pupal stage. Most pupae shifted from 22° C to 29° C undergo metamorphosis and eclose as adults. Adults are viable if placed at 29° C; however, all females and some males become sterile if maintained at this temperature.Somatic recombination was used to induce clones homozygous for a null allele of Ubl at different stages of development. Clones of this null allele appear to be cell lethal indicating that the Ubl ⁺ gene product is required at all stages of development. The viability of Ubl ^ts pupae and adults at 29° C may result from only a partial reduction in activity caused by the mutation at this nonpermissive temperature.     

Topology of the caudal fat body of the tumor-w mutant of Drosophila melanogaster

Hereditary melanotic tumors in the tumor^w strain of Drosophila melanogaster are known to involve encapsulation of the caudal fat body by the larval hemocytes. The encapsulated masses are subsequently melanized. The present study shows that the chain of events preceding encapsulation includes disintegration of the basement membrane of the caudal fat body and the appearance of particulate materials between and around the dissociating fat cells.     

The temperature-sensitive mutation vir ts(virilizer) identifies a new gene involved in sex determination of Drosophila

Summary When XX animals homozygous for the temperature-sensitive mutation vir ^tsof virilizer (2–103.9) are raised at the restrictive temperature of 29° C, they are transformed into sterile intersexes with a morphology comparable to XX flies mutant at the sex-determining gene doublesex (dsx). The gonads of the vir ^tsintersexes are ovaries in which the germ cells undergo abortive oogenesis. At the permissive temperature of 25° C or below, XX vir ^tsanimals develop into marginally fertile females. The temperature-sensitive period of vir ^tsis within the third larval instar. XY males are not affected by the mutation. Animals that are homozygous for vir ^tsand either transformer (tra) or tra2 develop as pseudomales; on the other hand, constitutive expression of a female-specific tra product rescues XX animals from the effect of vir ^ts, but these females are sterile. The data show that tra and tra2 are epistatic to vir. Animals with only one wildtype copy of either tra or tra2 and mutant for vir ^tsare already transformed into intersexes at 25° C. Conversely, the presence of three copies of the tra ⁺ gene largely prevents the effect of vir ^tsat 29° C; such flies are practically female, but sterile. Animals homozygous for vir ^tsand heterozygous for dsx ^D/+, raised at 29° C, are transformed into severely masculinized intersexes or almost pseudomales. The observations suggest that vir acts above and via tra and tra2 to achieve proper female-specific expression of the dsx gene in XX zygotes. Offprint requests to: R. Nöthiger     

Growth rates of North Sea macroalgae in relation to temperature,irradiance and photoperiod

Three eulittoral algae(Ulva lactuca, Porphyra umbilicalis, Chondrus crispus) and one sublittoral alga(Laminaria saccharina) from Helgoland (North Sea) were cultivated in a flow-through system at different temperatures, irradiances and daylengths. In regard to temperature there was a broad optimum at 10–15° C, except inP. umbilicalis, which grew fastest at 10 °C. A growth peak at this temperature was also found in four of 17 other North Sea macroalgae, for which the growth/temperature response was studied, whereas 13 of these species exhibited a growth optimum at 15 °C, or a broad optimum at 10–15 °C. Growth was light-saturated inU. lactuca, L. saccharina andC. crispus at photon flux densities above 70 µE m^–2s^–1, but inP. umbilicalis above 30 µE m^–2s^–1. Growth rate did not decrease notably in the eulittoral species after one week in relatively strong light (250 µE m^–2s^–1), but by about 50 % in the case of the sublittoralL. saccharina, as compared with growth under weak light conditions (30 µE m^–2s^–1). In contrast, chlorophyll content decreased in the sublittoral as well as in the eulittoral species, and the greatest change in pigment content occurred in the range 30–70 µE m^–2s^–1. Growth rate increased continuously up to photoperiods of 24 h light per day inL. saccharina andC. crispus, whereas daylength saturation occurred at photoperiods of more than 16 h light per day inU. lactuca andP. umbilicalis.     

Isolation and characterization ofgrandchildless-like mutants inDrosophila melanogaster

Summary Two temperature-sensitive sex-linkedgrandchildless (gs)-like mutations (gs(1)N26 andgs(1)N441) were induced by ethylmethane sulphonate inDrosophila melanogaster. They complemented each other and mapped at two different loci (1–33.8±0.7 forgs(1)N26 and 1–39.6±1.7 forgs(1)N441), which were not identical to those of any of thegs-like mutants reported in earlier work.Homozygous females of the newly isolated mutants produced eggs that were unable to form pole cells and developed into agametic adults. Competence of the embryos to form pole cells was not restored by wild-type sperm in either mutant; that is, the sterility caused by these mutations is controlled by a maternal effect.Fecundity and fertility ofgs(1)N26 females were low, and their male offspring showed a higher mortality than that of female offspring, causing an abnormal sex ratio. The frequency of agametic progeny was 93.1% and 55.8%, when the female parents were reared at 25° C and 18° C, respectively. In eggs produced by thegs(1)N26 females reared at 25° C, the migration of nuclei to the posterior pole was abnormal, and almost no pole cell formation occurred in these egg. Furthermore, half of these eggs failed to cellularize at the posterior pole. When the females were reared at 18° C, almost all of the eggs underwent complete blastoderm formation, and in half of these blastoderm embryos normal pole cells were formed.In the other mutant,gs(1)N441, the fecundity and fertility of the females were normal. The agametic frequency in the progeny was 70.8% and 18.6% when the female parents were reared at 25° C and 18° C, respectively. In the eggs laid by females reared either at 25° C or at 18° C, the migration of nuclei to the periphery and cellularization proceeded normally; nevertheless, in the majority of the embryos no pole cell formation occured at the stage when nuclei penetrated into the periplasm. When the females were reared at 18° C, some of the embryos from these females formed some round blastoderm cells with cytologically recognizable polar granules and nuclear bodies, which are attributes of pole cells. The temperature sensitive period ofgs(1)N441 was estimated to extend from stage 9 to 13 of King's stages of oogenesis.     

Respiratory studies on two small freshwater amoebae

The changes in respiration rate and mean cell volume induced by temperature within the range 10°C–25°C were investigated in two small species of freshwater amoebae,Saccamoeba limax Page andVannella sp. Mean cell volume varied in response to temperature, with maxima at 20°C inVannella sp. (10.15× 10³ (±1.80)m³ and 15°C inS. limax (9.08×10³ (±0.93)m³. Respiration rate increased over the temperature range investigated. The highest rates and the greatest rate of increase between temperatures occurred inVannella sp. Q₁₀ ranged between 0.12 and 1.33 inS. limax and between 1.77 and 7.36 inVannella sp. A regression of log oxygen uptake versus log cell volume incorporating the data of the present investigation and the data of other workers on amoeba respiration is presented, and the ecological significance and application of such data discussed.     

Erythromycin resistant mutations in Bacillus subtilis cause temperature sensitive sporulation.

Summary All of several hundred erythromycin resistant (ery^R) single site mutants ofBacillus subtilis W168 are temperature sensitive for sporulation (spo^ts). The mutants and wild type cells grow vegetatively at essentially the same rates at both permissive (30° C) and nonpermissive (47° C) temperatures. In addition, cellular protein synthesis, cell mass increases and cell viabilities are similar in mutant and wild type strains for several hours after the end of vegetative growth (47° C). In the mutants examined, the temperature sensitive periods begin when the sporulation process is approximately 40% completed, and end when the process is 90% complete. At nonpermissive temperatures, the mutants produce serine and metal proteases at 50% of the wild type rate, accumulate serine esterase at 16% of the wild type rate, and do not demonstrate a sporulation related increase in alkaline phosphatase activity.The ery^R and spo^ts phenotypes cotransform 100%, and cotransduce 100% using phage PBS1. Revertants selected for ability to sporulate normally at 47° C (spo⁺), simultaneously regain parental sensitivity to erythromycin. No second site revertants are found.Ribosomes from ery^R spo^ts strains bind erythromycin at less than 1% of the wild type rate. A single 50S protein (L17) from mutant ribosomes shows an altered electrophoretic mobility. Ribosomes from spo⁺ revertants bind erythromycin like parental ribosomes and their proteins are electrophoretically identical to wild type. These data indicate that the L17 protein of the 50S ribosomal subunit fromBacillus subtilis may participate specifically in the sporulation process.     

A temperature effect on nondisjunction of the X chromosomes among eggs from agedDrosophila females

The effects of temperature and aging on the frequency of nondisjunction inDrosophila melanogaster eggs were investigated. At 25°C offspring arising from 3–5 day old control females had a nondisjunction frequency (0.943/1000 offspring) very similar to that for females who were 24–26 or 27 days old when eggs were collected (1.044/1000 offspring). When females were aged for the same length of time at 10°C the frequency of nondisjunctional exceptions increased to 3.368 per 1000 offspring. These results indicate that aging the females at 25°C does not increase the nondisjunction rate over that obtained from non-aged females raised at 25°. The increase in nondisjunction frequencies when the females were aged at 10°C reflects an influence of temperature on the meiotic process inDrosophila melanogaster. At the low temperature eggs were also aged since few or no eggs were laid during the aging process. Thus in addition to a temperature effect on nondisjunction rates at 10°C there may also be an age effect.     

Polyphenolic Antioxidants Efficiently Protect Urease from Inactivation by Ultrasonic Cavitation

Inactivation of urease (25 nM) in aqueous solutions (pH 5.0–6.0) treated with low-frequency ultrasound (LFUS; 27 kHz, 60 W/cm², 36–56°C) or high-frequency ultrasound (HFUS; 2.64 MHz, 1 W/cm², 36 or 56°C) has been characterized quantitatively, using first-order rate constants: k _in, total inactivation; k _in ^*, thermal inactivation; and k _in(us), ultrasonic inactivation. Within the range from 1 nM to 10 M, propyl gallate (PG) decreases by approximately threefold the rate of LFUS-induced inactivation of urease (56°C), whereas resorcinol poly-2-disulfide stops this process at 1 nM or higher concentrations. PG completely inhibits HFUS-induced inactivation of urease at 1 nM (36°C) or 10 nM (56°C). At 0.2–1.0 M, human serum albumin (HSA) increases the resistance of urease treated with HFUS to temperature- and cavitation-induced inactivation. Complexes of gallic acid polydisulfide (GAPDS) with HSA (GAPDS–HSA), formed by conjugation of 1.0 nM GAPDS with 0.33 nM HSA, prevent HFUS-induced urease inactivation (56°C).     

The half-life of mRNA in Saccharomyces cerevisiae.

Summary The decay kinetics of mRNA was studied in a yeast temperature-sensitive mutant, ts136, which is defective in cytoplasmic RNA production at 37° C. The disappearance of the synthetic capacity of mRNA was determined by withdrawing equal volumes of ts136 cell culture and pulse-labelling with [³⁵S]methionine at various time intervals after the shift to 37° C from 23° C. The synthesized proteins were separated on a two-dimensional gel electrophoretic system and then quantitatively analyzed for their incorporated radioactivities by scintillation counting. Our results show that yeast mRNAs have divergent functional half-lives ranging from 4.5 to 41 min, with an average value of 22 min. Each mRNA exhibits a simple exponential decay with its own characteristic decay pattern. Of the approximately 500 major polypeptides made by yeast cells, which are detectable on autoradiograms of the gels, 80 were arbitrarily selected and the mRNAs coding for those polypeptides were examined for their decay kinetics.     

Genes involved in the regulation of the neutral phosphatase in Chlamydomonas reinhardi

Summary In Chlamydomonas reinhardi, mutations in either of two unlinked genes (PD₂ and PD₃) abolish the activity of the derepressible neutral phosphatase. The question arose whether these genes (or one of them) specify the structure of the enzyme or whether they have a regulatory function.Three mutants producing an active phosphatase at 25°C but not at 35°C were isolated and investigated. One of these mutants (PD ₁₁ ^ts ) was allelic with PD₂, another one (PD ₁₂ ^ts ) was linked to PD₃ and the third one (PD ₁₃ ^ts ) was linked to PD₂.PD ₁₁ ^ts and PD ₁₃ ^ts affected the formation of the neutral phosphatase only whereas PD ₁₂ ^ts interfered with the formation of both neutral and alkaline phosphatases at 35°C. The neutral phosphatase produced by the three mutants at low temperature was not more thermosensitive in vitro than the wild enzyme. Moreover, quite similar K_m values were found in WT, PD ₁₁ ^ts and PD ₁₂ ^ts using naphthyl phosphate as a substrate.On the other hand, revertants of PD ₂ ^- and PD ₃ ^- were isolated: their neutral phosphatases could not be distinguished from the wild enzyme on the basis of their thermosensitivities and K_m values for naphthyl phosphate.These results are consistent with the idea that PD₂ and PD₃ are regulatory genes. Other possible regulatory genes were revealed through PD ₁₂ ^ts and PD ₁₃ ^ts mutations.Chercheur qualifié du Fonds National Belge de la Recherche Scientifique     

High efficiency temperature-sensitive amber suppressor strains of Escherichia coli K12

Summary A set of eight strains combining the supD43,74 ts¹ suppressor gene with alleles of three suppressor-enhancing (sue) genes have been constructed and characterized. The sue mutations work cooperatively to raise suppressor activity and together raise the activity of the supD43, 74-encoded suppressor 40-fold. These strains further expand the utility of the ts suppressor system by providing as much as 100% suppressor activity at temperatures at or below 20°C to as little as 0.015% suppressor activity at 43°C.     

Physiological suppression of the temperature-sensitive sporulation defect in a Bacillus subtilis RNA polymerase mutant

Summary Five hundred putative RNA polymerase mutants of Bacillus subtilis were isolated by selecting for resistance to the RNA polymerase inhibitors rifampin (Rif^r), streptovaricin (Str^r) or streptolydigan (Std^r). This collection was screened for mutants that were unable to sporulate at the non-permissive temperature of 46°C, yet which sporulated well at 37°C and had normal vegetative growth (Spo^ts phenotype). Nearly one half of the Rif^r and one quarter of the Stv^r mutants were Spo^ts, whereas none of the Std^r mutants had this phenotype.The streptovaricin resistant strain stv84 was studied in detail. The stv84 mutation maps between cysA14 and strA39 on the B. subtilis chromosome, and the Stv^r and Spo^ts phenotypes cotransform at a frequency of 100%. The Spo^ts phenotype of stv84 could be physiologically corrected by supplementing the growth medium with inhibitors of RNA synthesis such as rifampin or azauracil, with carbohydrates such as ribose, mannose or glycerol, or with lipids such as Tween 40 or fatty acids native to Bacillus subtilis membranes. A Spo^ts phenotype resembling that of stv84 was produced in wild type B. subtilis by adding cerulenin, an inhibitor of fatty acid biosynthesis, to the growth medium. This cerulenin-induced sporulation defect was reversed by the same treatments that correct the temperature-sensitive genetic defect of stv84. These data indicate that the Spo^ts phenotype of strain stv84 is not due to an intrinsic inability of the mutant RNA polymerase to transcribe developmentally-specific genes at the nonpermissive temperature. Rather, the data suggest that the stv84 lesion causes a physiological imbalance which disrupts membrane structure or function in sporulating cells.