Mariann Bienz

A male accessory gland peptide that regulates reproductive behavior of female D. melanogaster

The adult male accessory glands of D. melanogaster synthesize and secrete a peptide that represses female sexual receptivity and stimulates oviposition. Normally, this peptide is transferred to females during copulation; however, the peptide shows the same biological activity after purification and subsequent injection into the abdominal cavity of female virgins. Amino acid sequencing of the purified peptide and oligonucleotide-directed cDNA cloning established that the peptide consists of 36 amino acids. It appears to be synthesized as a precursor with a hydrophobic signal sequence of 19 residues at its N-terminal end. The precursor peptide is encoded by a short mRNA that accumulates exclusively in the male accessory gland. The gene has been localized by in situ hybridization to polytene chromosomes at 70A.

The APC tumour suppressor has a nuclear export function.

The adenomatous polpyposis coli (APC) protein is mutated in most colorectal tumours. Nearly all APC mutations are truncations, and many of these terminate in the mutation cluster region located halfway through the protein. In cancer cells expressing mutant APC, β-catenin is stabilized and translocates into the nucleus to act as a transcriptional co-activator of T-cell factor. During normal development, APC also promotes the destabilization of β-catenin and Drosophila Armadillo. It does so by binding to the Axin complex which earmarks β-catenin/Armadillo for degradation by the proteasome pathway. APC has a regulatory role in this process, which is poorly understood. Here we show that APC contains highly conserved nuclear export signals 3′ adjacent to the mutation cluster region that enable it to exit from the nucleus. This ability is lost in APC mutant cancer cells, and we provide evidence that β-catenin accumulates in the nucleus as a result. Thus, the ability of APC to exit from the nucleus appears to be critical for its tumour suppressor function.

Induction across germ layers in Drosophila mediated by a genetic cascade

We report an induction process occurring between two germ layers in the Drosophila embryo that involves a cascade of five interacting genes. Two of these, Ultrabithorax and abdominal-A, encode nuclear homeobox proteins; each of them is expressed in one of two adjacent parasegments in the visceral mesoderm and directs expression in its parasegment of a separate target gene, decapentaplegic in parasegment 7 and wingless in parasegment 8. The activity of both target genes is required for normal expression of another homeotic gene, labial, in cells of the adhering midgut epithelium. Their products are putative extracellular proteins, which presumably act as signals between the two germ layers. Positional instruction of this kind may be needed since the endoderm, unlike the mesoderm, appears unsegmented at first as it originates from two primordia near the embryonic poles, outside the realm of segmentation genes.

Mechanisms of Heat-Shock Gene Activation in Higher Eukaryotes

Publisher Summary This chapter discusses the components of heat-shock gene-activation systems: the cis -acting elements and the trans -acting factors. The chapter also explains the way these components act together to result in transcription activation and the way multiple controls are achieved. It also illustrates the way a cell detects the environmental stimulus and translates it into gene activation and the way the functions of gene products relate to this process. The chapter focuses on heat-shock gene activation in higher eukaryotes and relates this to the field of higher eukaryotic transcription. Heat-shock gene induction represents a model case for gene activation in two respects. During transient heat induction, heat-shock genes undergo reprogramming from the off- to the on-state; heat-shocked and unshocked cells differ in their states of gene activation in a way analogous to two different cell types or cell lineages. Some heat-shock genes are not only transiently heat inducible but also activated during normal cell growth or development, which leads to the question of how multiple control mechanisms act on the same gene.

Wild-type tRNATyrG reads the TMV RNA stop codon, but Q base-modified tRNATyrQ does not

Although protein synthesis usually terminates when a stop codon is reached along the messenger RNA sequence, there are examples, mainly in viruses, of the stop codon being suppressed by a tRNA species. A strong candidate for this phenomenon occurs in tobacco mosaic virus (TMV) in the form of two proteins (110K and 160K, of molecular weights 110,000 and 160,000, respectively)1, sharing an N-terminus sequence, which are translated in vitro from a purified species of viral RNA. We have investigated the identity of the tRNA responsible for production of the 160K protein and show here that it is one of the tyrosine tRNAs. Another tyrosine tRNA, in which the first base of the anticodon is highly modified, does not act as a suppressor, indicating the possible regulatory function of such modifications.

Differential regulation of Ultrabithorax in two germ layers of drosophila

The homeotic gene Ultrabithorax (Ubx) is expressed in specific parts of Drosophila embryos: in a single metamer in the visceral mesoderm and forming a complex pattern limited to a broad domain in the ectoderm and in the somatic mesoderm. Here we use a linked beta-galactosidase gene to identify cis-acting regulatory sequences. In the visceral mesoderm, correct expression of Ubx depends on localized upstream sequences. In the ectoderm, all galactosidase-positive transformants show the same characteristic pattern. The repeated elements of this basal pattern appear to be a sub-pattern of engrailed (en) expression; they depend on en function as well as on sequences in the Ubx RNA leader. We use a mutant (Haltere-mimic) to show that sequences that normally restrict segmental expression of Ubx in the ectoderm are located downstream from the RNA leader.

Evidence that the Abdominal-B r element function is conferred by a trans-regulatory homeoprotein.

The Abdominal‐B gene is a homeotic gene located in the distal‐most region of the bithorax complex (BX‐C). Based on complementation analysis it has been proposed that the gene contains two separable genetic elements, called the m and r elements. The r element has a chiefly regulatory function confined to parasegment 14. In a reverse Northern screen of the distal‐most BX‐C DNA, we found four distinct areas that are transcribed in the embryo. One of the transcripts spans a large genomic region which, upon disruption by rearrangement breakpoints, causes loss of r element function. This transcript is expressed in the early embryo in a single domain apparently corresponding to parasegment 14. We propose that the small homeoprotein encoded by the transcript acts as a trans‐regulator to confer r element function.

Coordinate action of a proximal homeoprotein binding site and a distal sequence confers the Ultrabithorax expression pattern in the visceral mesoderm

Spatially regulated expression of the homeotic gene Ultrabithorax (Ubx) in the visceral mesoderm can be mimicked in transformed Drosophila embryos by expression of a Ubx‐‐beta‐galactosidase fusion gene. Here we show that a proximal homeoprotien binding sequence downstream of the Ubx transcription start site, the B element, is required for this pattern. A distal upstream Ubx sequence, but not the B element, is sufficient to confer the pattern if linked to an hsp70 TATA box in a heterologous construct. The pattern in this case requires Ubx function, like endogenous Ubx expression in the visceral mesoderm, suggesting that the distal upstream sequence contains an important target sequence for autoregulation. We propose that the B element, in the context of the Ubx promoter, functions to mediate enhancer function of the distal sequence. Thus, the visceral mesoderm pattern requires coordinate action of a proximal and a distal regulatory element.

The structure of the Ultrabithorax promoter of Drosophila melanogaster.

The sequence of 4118 nucleotides upstream of the putative initiator codon of the D. melanogaster Ultrabithorax protein was determined. The transcription initiation site for the corresponding mRNA was identified by S1 nuclease mapping and primed extension. It appears that all embryonic RNA products that encode the first protein exon are initiated approximately one kilobase upstream of the initiator methionine, suggesting a unique Ultrabithorax promoter. The unusually long mRNA leader is unspliced. Upstream of the initiator codon occur two additional methonine codons; the first one is followed by an open reading frame encoding a putative polypeptide of 69 amino acids. We discuss the role of the leader and 5′ flanking sequences with respect to transcriptional and posttranscriptional control of Ultrabithorax gene expression.

Functional dissection of Drosophila abdominal-B protein.

The Abdominal-B gene is unique among homeotic Drosophila genes as it encodes two proteins m and r which confer different functions. The m protein corresponds to an r protein with a large N-terminal extension; the two proteins contain the same homeodomain. We have used a transient co-transfection assay, based on HeLa cells, to analyse the intrinsic function of m and r protein in activating transcription. We find two strong transcriptional activation domains in the common part of the two proteins. The m-specific exon contains additional transcriptional activation potential. Despite this, the m protein is a weaker transcriptional activator than the r protein. Apparently, there are inhibitory sequences in the m-specific exon which, in the embryo, may have a role in masking r function in the intact m protein.