A chromosome walk linking the gatA and alcC genes of Aspergillus nidulans

The amdA gene of Aspergillus nidulans has been mapped to linkage group VII, between the gatA and alcC genes (Jones and Sealy- Lewis 1990 Curr. Genet. 17:81-85). As clones of both of the genes flanking amdA were available, they were used as starting points for chromosome walking towards amdA. The relative orientation of these genes on linkage group VII was not known at the time. Therefore, both walks extended in both directions until overlapping clones from the different walks were detected. The total distance covered by all walks is approximately 240 kb

Multiple doublesex-Related Genes Specify Critical Cell Fates in a C. elegans Male Neural Circuit

In most animal species, males and females exhibit differences in behavior and morphology that relate to their respective roles in reproduction. DM (Doublesex/MAB-3) domain transcription factors are phylogenetically conserved regulators of sexual development. They are thought to establish sexual traits by sex-specifically modifying the activity of general developmental programs. However, there are few examples where the details of these interactions are known, particularly in the nervous system.In this study, we show that two C. elegans DM domain genes, dmd-3 and mab-23, regulate sensory and muscle cell development in a male neural circuit required for mating. Using genetic approaches, we show that in the circuit sensory neurons, dmd-3 and mab-23 establish the correct pattern of dopaminergic (DA) and cholinergic (ACh) fate. We find that the ETS-domain transcription factor gene ast-1, a non-sex-specific, phylogenetically conserved activator of dopamine biosynthesis gene transcription, is broadly expressed in the circuit sensory neuron population. However, dmd-3 and mab-23 repress its activity in most cells, promoting ACh fate instead. A subset of neurons, preferentially exposed to a TGF-beta ligand, escape this repression because signal transduction pathway activity in these cells blocks dmd-3/mab-23 function, allowing DA fate to be established. Through optogenetic and pharmacological approaches, we show that the sensory and muscle cell characteristics controlled by dmd-3 and mab-23 are crucial for circuit function.In the C. elegans male, DM domain genes dmd-3 and mab-23 regulate expression of cell sub-type characteristics that are critical for mating success. In particular, these factors limit the number of DA neurons in the male nervous system by sex-specifically regulating a phylogenetically conserved dopamine biosynthesis gene transcription factor. Homologous interactions between vertebrate counterparts could regulate sex differences in neuron sub-type populations in the brain

A Cholinergic-Regulated Circuit Coordinates the Maintenance and Bi-Stable States of a Sensory-Motor Behavior during Caenorhabditis elegans Male Copulation

Penetration of a male copulatory organ into a suitable mate is a conserved and necessary behavioral step for most terrestrial matings; however, the detailed molecular and cellular mechanisms for this distinct social interaction have not been elucidated in any animal. During mating, the Caenorhabditis elegans male cloaca is maintained over the hermaphrodite's vulva as he attempts to insert his copulatory spicules. Rhythmic spicule thrusts cease when insertion is sensed. Circuit components consisting of sensory/motor neurons and sex muscles for these steps have been previously identified, but it was unclear how their outputs are integrated to generate a coordinated behavior pattern. Here, we show that cholinergic signaling between the cloacal sensory/motor neurons and the posterior sex muscles sustains genital contact between the sexes. Simultaneously, via gap junctions, signaling from these muscles is transmitted to the spicule muscles, thus coupling repeated spicule thrusts with vulval contact. To transit from rhythmic to sustained muscle contraction during penetration, the SPC sensory-motor neurons integrate the signal of spicule's position in the vulva with inputs from the hook and cloacal sensilla. The UNC-103 K+ channel maintains a high excitability threshold in the circuit, so that sustained spicule muscle contraction is not stimulated by fewer inputs. We demonstrate that coordination of sensory inputs and motor outputs used to initiate, maintain, self-monitor, and complete an innate behavior is accomplished via the coupling of a few circuit components

Cloning and characterisation of the regulatory gene, amdA, of Aspergillus nidulans

TypescriptThesis (PhD )-- University of Melbourne, Faculty of Science, 1994Includes bibliographical references (leaves 160-177)Semi-dominant mutations in the amdA gene lead to elevated expression of the gene encoding acetamidase, amdS. These mutations also cause constitutive expression of the acetate-inducible gene, aciA. In the amdS 5' regulatory region, two cis-acting mutations, amdI66 and amdI666, have been isolated which specifically affect amdA activation of amdS. These mutations affect an 18 bp GA-rich sequence, thought to define the amdA site of action within the amdS promoter region. Similar GA-rich sequences have also been found in the 5' region of aciA. The work presented in this thesis describes the cloning and initial functional characterisation of the amdA gene and three of its mutant alleles. The wild-type amdA gene has been cloned by undertaking a chromosome walk from genes gatA and alcC on linkage group VII and localised by complementation of an amdA loss-of-function mutation. Transcriptional analysis reveals that the gene is expressed at low levels and constitutively under growth conditions which affect expression of amdS and aciA. The gene is predicted to encode an 880 amino acid protein which contains two C2H2 zinc fingers, similar to those found in Xenopus laevis transcription factor TFIIIA, a potential nuclear localisation sequence and two potential transcriptional activation domains. Functional analysis of in vitro-generated deletion mutations and of the semi-dominant mutant allele, amdA7, indicate that the acidic activation domain identified in the protein sequence is functionally significant. Interestingly, the C2H2 zinc finger motifs identified in the protein are similar to those found in the carbon catabolite repressor protein, CreA, which also regulates amdS and recognises sequences which coincide with the proposed site of action for AmdA within the promoter of this structural gene

Regulation of sex-specific differentiation and mating behavior in C. elegans by a new member of the DM domain transcription factor family

Mutations in Caenorhabditis elegans gene mab-23 cause abnormal male tail morphology and abolish male fecundity but have no obvious effect in the hermaphrodite. Here we show that mab-23 encodes a DM (Doublesex/MAB-3) domain transcription factor necessary for specific aspects of differentiation in sex-specific tissues of the male. mab-23 is required for the patterning of posterior sensory neurons in the male nervous system, sex muscle differentiation, and morphogenesis of the posterior hypodermis, spicules, and proctodeum. Failure of mab-23 mutant males to sire progeny is due primarily to defective sex muscle-mediated turning during copulatory behavior and likely compounded by impairment of sperm passage through the proctodeum. In the male nervous system, mab-23 refines ray neuron subtype distribution by restricting expression of dopaminergic neurotransmitter identity through interactions with the Hox gene egl-5 and a TGF-β-related signaling pathway. mab-23 has distinct roles and functions independent of mab-3, indicating different aspects of C. elegans male sexual differentiation are coordinated among DM domain family members. Our results support the hypothesis that DM domain genes derive from an ancestral male sexual regulator and suggest how regulation of sexual development has evolved in distinct ways in different phyla