摘要:

AbstractSeveral genes that are deactivated upon the initiation of development ofDictyostelium discoideumhave been identified by differential screening of various cDNA libraries. These genes have in common a decrease in the steady‐state levels of their corresponding mRNAs as development proceeds. When development was carried out in the absence of protein synthesis by inhibition with cycloheximide, the decrease in mRNA levels for most genes (V genes) was normal or slightly accelerated. However, for about 5% of the genes (H genes), cycloheximide caused an apparent induction of expression, as revealed by a slight or dramatic increase in mRNA levels instead of the normal decrease. This effect was due to inhibition of protein synthesis and not to cycloheximide per se. The induction was found to be due to an enhancement of the trascription rate; normal rates of transcription for the H genes were dependent upon continued protein synthesis during vegetative growth and during development. Thus, two general regulatory classes exist for deactivation of gene expression upon initiation of development, one dependent and one independent of protein synthesis. Models concerning the control of expression of these two classes of genes are discussed here. Analysis of expression of these genes in mutant strains that are aggregation‐deficient has also been performed, and the results lead to subdivisions of the clas

ISSN:0192-253X    

DOI:10.1002/dvg.1020090414

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractWe have examined the expression of a cAMP pulse‐repressed and two cAMP pulse‐induced genes in response to cAMP and caffeine under a number of different physiological conditions, and in several classes of developmental mutants altered in cAMP‐mediated signal transduction pathways. The data presented help characterize the mutants with regard to early gene expression. Analysis of the data indicates that full induction of the pulse‐induced or repression of the pulse‐repressed genes requires cycles of activation and adaptation of the cAMP receptor but does not require a rise in intracellular cAMP. Comparison of the results obtained between different mutant classes suggests that repression and activation of the two classes of genes can be uncoupled, implying that different intracellular mechanisms control these processes. In addition, we examined the effects of caffeine and show that it can induce pulse‐induced mRNA accumulation in the abse

ISSN:0192-253X    

DOI:10.1002/dvg.1020090415

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractInDictyostelium, extracellular cAMP interacts specifically with cell‐surface receptors to promote the accumulation of a variety of intracellular second messengers, such as, 3′‐5′ cyclic adenosine monophosphate (cAMP) and 1,4,5 inositol trisphosphate (IP3). We and others have shown that activation of the cell‐surface cAMP receptor can also modulate the expression of theDictyosteliumgenome during development. In at least one instance, synthesis of intracellular cAMP is required for appropriate gene regulation. However, the induction of most cAMP‐dependent gene expression can occur in the absence of receptor‐mediated activation of adenylate cyclase and a consequent accumulation of intracellular cAMP. These results suggest that other intracellular second messengers produced in response to receptor activation may potentially act as signal transducers to modulate gene expression during development. In vertebrate cells, IP3and diacylglycerol (DAG) are intracellular activators of specific protein kinases; they are produced in equimolar amounts by cleavage of phosphoinositol bisphosphate after a receptor‐mediated activation of a membrane‐bound phosphodiesterase. IP3and, thus, by inference, diacylglycerol are synthesized inDictyosteliumas a response to cAMP interacting with its cell‐surface receptor. Using defined conditions to inhibit the accumulation of extracellular cAMP, we have examined the effects of these compounds on the accumulation of extracellular cAMP, we have examined the effects of these compounds on the expression of genes that require cAMP for their maximal expression. Our results suggest that intracellular IP3and DAG may in part mediate the action of extracellular cAMP on the expression of the

ISSN:0192-253X    

DOI:10.1002/dvg.1020090416

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractElevated levels of cAMP are essential for the expression of many postaggregation prespore and prestalk mRNA species and for the suppression of some growth phase mRNAs. Here we review evidence that this regulation is mediated by cAMP interacting at the cell surface receptor. These effects of cAMP on gene expression can occur under conditions where the receptor‐associated adenylate cyclase is inactivated and in concentrations that are consistent with receptor binding. A number of differences are noted in the mechanism by which cAMP regulates prespore and prestalk genes. Finally, evidence is reviewed for the role of a Ca2+‐dependent signal transduction system in coupling the expression of some of the prespore mRNAs to the cAMP receptor. This signal transduction system does not appear to be involved in the expression of the cAMP‐dependent prestalk

ISSN:0192-253X    

DOI:10.1002/dvg.1020090417

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractcAMP regulates gene expression inDictyostelium discoideumthrough the cell surface receptor and is therefore a transmembrane signal transduction event. We have now begun to examine the signal transduction pathway that transmits the cAMP‐induced signal to the nucleus. The results presented here indicate that Ca2+plays a crucial role. A comparison of the accumulation of UDPGP1 mRNA during development with the corresponding transcription rates revealed that this gene is regulated primarily at the level of transcription. To elucidate the factors involved in the regulation of the UDPGP1 gene we characterized itscisacting sequences. We constructed a series of deletions into the 5′ flanking region of the UDPGP1 gene and analyzed the expression of the mutated DNA in transformants. A sequence element essential for the expression of the UDPGP1 gene is located between ‐500 bp and ‐288 dp from the transcription start site. This promoter element appears to be a short G + C‐rich sequence positioned between ‐374 to ‐395 and coincides with a DNase I hypers

ISSN:0192-253X    

DOI:10.1002/dvg.1020090418

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractThe nucleotide sequence of D19, aDictyosteliumgene that encodes a prespore‐specific mRNA sequence shows it to encode PsA, the cell surface protein detected by the MUD 1 monoclonal antibody. The predicted sequence of the protein reveals a largely hydrophobic C terminus, with chemical similarity to proteins known to be attached to the plasma membrane via a phosphatidylinositol link. The C‐terminal region has direct sequence homology to the contact sites A protein and to the phosphatidylinositol‐linked form of a chicken N‐CAM, suggesting that it might play a role in cell adhesion. Expression of the D19 gene is known to be induced by cAMP and repressed by adenosine. The accumulation of the D19 mRNA is also repressed by DIF, the putative stalk‐specific morphogen, and this effect is mediated at the transcriptional level. The pDd56 and pDd63 genes are induced by DIF, and they are specific markers of prestalk and stalk cells. They encode, respectively, ST310 and ST430, two proteins that were first identified by two‐dimensional gel electrophoresis. Both proteins are predominantly composed of a highly conserved, 24‐amino acid repeat. The two proteins are localized in the slime sheath of the migratory slug and in the stalk tube and stalk cell wall of the mature culminant, where they presumably function as structural components of the extracellular matrix. We have constructed marked derivatives of the pDd56, pDd63, and D19 genes, and these are correctly regulated after transformation intoDictyosteliumcells. Thus we have determined the structure, and elucidated possible functions, for one prespore and two prestalk genes. These sequences should be of value, both as markers of the earliest events in cellular differentiation and in identifying the regulatory sequences controlling cell type‐specific

ISSN:0192-253X    

DOI:10.1002/dvg.1020090419

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractThis paper reviews our studies of three aspects of post‐transcriptional regulation inDictyostelium discoideum:(1) the determinants of mRNA stability in vegetative amoebae; (2) the effects of disaggregation and cyclic AMP on the decay rates of cell‐type‐specific mRNAs in late developing cells; and (3) the cytoplasmic function of the 3′ poly(A) tracts present on most mRNAs. We find that: (1) mRNA stability in vegetative amoebae is not dependent on mRNA size, ribosome loading, or poly(A) tract length, but may be determined by specific 3′‐untranslated sequences within a given mRNA; (2) mRNA decay rates in late developing cells are heterogeneous, and cyclic AMP does not act directly to stabilize cell‐type‐specific mRNAs; and (3) poly(A) is most likely involved in the initiation of protein synthesis via an interaction with cytoplasmic poly(A)

ISSN:0192-253X    

DOI:10.1002/dvg.1020090420

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractWe have isolated recombinant plasmids that contain cDNA inserts complementary to mRNAs encoding six different r‐proteins ofDictyostelium discoideum.Southern and quantitative dot blot analyses have shown that each of the r‐protein genes represented in these plasmids is encoded by a single copy gene and that these genes are not tightly linked to each other. We have determined the relative amount of the six r‐protein mRNAs present in cells at intervals throughout development and find that for the first 9 hours of development, each of the mRNAs remains present at virtually the same level as in vegetatively growing cells. Between 9 and 11 hours of development, there is a rapid loss of these mRNAs to 15% or less of vegetative levels, and that low level remains, or slightly declines, through the late stages of development. We have shown that two post‐transcriptional events contribute to the developmental regulation of the expression of the r‐protein genes. The first involves a specific block to translational initiation that is not the result of inactivation of these mRNAs by decapping or deadenylation. The second is a change in the stability of these mRNAs during early development. In order to begin to analyze the role of specific sequences that may act as targets or signals in these events, we have cloned and sequenced a 1.9‐kb genomic DNA fragment that encodes one of the r‐proteins. We find that transcription of this gene begins in a pyrimidine‐rich region that is not preceded by a TATA box, the gene contains a single intron of 350 bp, and there are two alternative 3′ processing sites. In addition, the 5′‐untranslated region of the transcript contains an unusually high percentage of G and C residues relative to ot

ISSN:0192-253X    

DOI:10.1002/dvg.1020090421

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractExpression of theDictyostelium discoideumpst‐cath (CP2) gene is transcriptionally regulated during multicellular development, and the gene is inducible in competent single cells following administration of exogenous cAMP. The 5′ flanking region of pst‐cath (CP2) that extends from −313 to the Cap site (+−1) has previously been shown to contain sufficientcis,‐acting regulatory elements for proper developmental and cAMP‐inducible expression of a foreign gene [Datta and Firtel, 1987, Mol Cell Biol 7:149–159]. The −283 to −201 region includes two exceptional “G‐boxes” centered at −233 and −217 respectively, and this ∼︁ 80 bp region is essential for basal as well as regulated expression of the pst‐cath (CP2) gene. Here we summarize results obtained from a detailed analysis of a series of linker‐scanner mutants and mutants that carry small internal deletions within the essential 80‐bp region. Insertion of a synthetic oligonucleotide that includes the downstream G‐box is demonstrated to rescue a low level of cAMP‐inducible expression following insertion into cassette mutants. The effect of introducing a change in the relative spacing between regulatory elements has also been investigated.We have analyzed nuclear extracts for the presence of DNA‐binding proteins that interact specifically with the pst‐cath (CP2) regulatory region and identified two such putativetrans‐acting factors: (1) the AT‐factor that is observed within a few hours following the onset of starvation and that binds tightly to stretches of alternating adenine‐thymine residues (poly(dA‐dT)); and (2) the AG‐factor that is present in nuclear extracts of aggregated cells. Competition studies have demonstrated significant differences in the affinity that characterizes the binding of the two factors to G‐box‐containing sequences. The binding specificities of these DNA‐binding proteins have been a

ISSN:0192-253X    

DOI:10.1002/dvg.1020090422

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY

摘要:

AbstractThe cysteine proteinase 1 (CP1) and cysteine proteinase 2 (CP2) genes ofDictyostelium discoideumencode coordinately expressed mRNA sequences that are inducible by extracellular cAMP. Both genes form part of divergently transcribed gene pairs. The gene proximal to CP1 is coordinately regulated and encodes a protein containing several potential zinc binding domains of the kind found in DNA binding proteins. The gene proximal to CP2 is a constitutively transcribed gene of unknown function. There are multiple, short, G‐rich sequence elements between both gene pairs, and deletion of the pair of elements 200 nucleotides upstream from the CP2 gene abolishes cAMP‐inducibility. A synthetic oligonucleotide, containing two copies of the G‐rich element from the CP1 gene, will reconstitute cAMP‐inducibility in the deletion mutant of the CP2 gene. This shows that the elements in the two genes are functionally homologous. Efficient induction requires at least two copies of the CP1 element, but their relative orientation is unimportant. Two copies in an inverted orientation are, however, inactive when moved upstream of their normal position and are incapable of conferring cAMP‐inducibility on a heterologous gene. These observations suggest that these sequences are either essential promoter elements, not themselves interacting with the inducer, or that their interaction with a separate class of control sequences is necessary for inducible e

ISSN:0192-253X    

DOI:10.1002/dvg.1020090423

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1988

数据来源: WILEY