InteractiveFly: GeneBrief

jim lovell: Biological Overview | References

The larvae of Drosophila melanogaster grow rapidly through use of a highly truncated cell cycle in which mitosis is entirely eliminated. The Drosophila homolog of the protooncogene transcription factor Myc plays a major role in promoting this endopolyploid (EP) growth. It was previously determined that the gene jim lovell (lov), which encodes a member of the BTB/POZ (Bric-a-brac, Tramtrack, Broad/Pox virus zinc finger) domain family of transcription factors, is also required for EP growth in one larval tissue, the trachea. This study shows that lov promotes EP growth in three further tissues indicating a fundamental role in this process. However, epistasis experiments revealed heterogeneity in lov's action in these tissues. Whereas in the tracheae and salivary glands lov acts downstream of Myc, in the fat body, reduced expression of lov does not impede the action of Myc, indicating an upstream action for the gene. This study shows that lov's regulation of the gene uninflatable (uif) in the tracheae is a component of this difference. uif is required for tracheal EP growth downstream of Myc and lov but has no equivalent role in the fat body. Although Uif is a transmembrane component of the plasma membrane in the tracheae, its action downstream of Myc suggests an intracellular role for the protein in the tracheae. In addition to regulating uif expression in some tissues it was also shown that lov locates to the nucleolus, indicating it can function in both polymerase I and polymerase II transcriptional events. The major finding of this study is that tissue-specific mechanisms can interact with universal growth promotion by Myc to generate the individual endopolyploid organs of the larvae (Zhou, 2020).

Within all multicellular organisms, mitotic cell cycling is the dominant mechanism for producing tissue and organismal growth. However, variant modes of growth exist in which supernumerary copies of the genome, contained in one or more nuclei, are generated within a single enlarged cytoplasm. These variant mechanisms are seen across evolution and have long been known to play roles in tissue differentiation. However, more recently some variants have been recognized as inducible growth phenomena activated by environmental stresses. In particular the ability of some cancer cells to resist apoptosis in response to drug-induced DNA damage involves transition to a polyploid state. Subsequently, a few cells can then revert to mitotic metastatic growth. In the variant growth mechanisms collectively termed endocycling (EC), rounds of genome replication occur without mitotic cytoplasmic division so that giant cells are generated. In the most extreme form of EC, here termed endopolyploid (EP) growth, the entire cell cycle M phase is absent, including nuclear envelope breakdown and chromosome segregation: the cell cycle consists solely of rounds of synchronized DNA synthesis (S) alternating with gap (G) phases. Thus in EP growth, giant cells with giant polyploid nuclei are generated (Zhou, 2020).

Giant cells synthesize fewer cell surface components than an equivalent mitotic cell mass. In addition, skipping elements of M phase saves both cellular resources and time. As a result, EC mechanisms are often associated with the need for rapid growth. EP growth, in which M phase is completely absent, offers the greatest potential for a rapid growth rate amongst the EC mechanisms. The major function of the larval stage in the Drosophila life cycle is that of extremely rapid growth. Over a four day period, larvae increase 200 fold in weight and their size at pupation defines that of the final adult. With the exception of a few tissue types (in particular, the imaginal discs and most of the nervous system) all of the larval tissues grow by an EP mechanism. The Drosophila larva thus offers an opportunity to examine the coordinated deployment of EP growth in multiple tissues and thus to determine whether differences in the regulation of EP growth exist in differing cell types (Zhou, 2020).

Although the signaling pathways that initiate EP growth in the larva are not well characterized, it is clear that the single Myc protein of Drosophila is a major downstream positive regulator of EP growth in the larval tissues (Pierce, 2004). Failed EP growth has been directly demonstrated in three Myc null larval tissues—salivary glands, fat body, and hindgut—and the essentially complete absence of growth in Myc null larvae argues for a regulatory role in all EP tissues (Pierce, 2004). Endoreplication of the nuclear DNA and overall cellular growth are tightly coordinated in EP growth. However, available evidence suggests that Myc does not directly affect DNA replication but rather acts to stimulate cellular growth by multiple genome-wide actions including increased ribosome synthesis, mRNA translation capacity and overall cellular metabolism. Myc is a bHLH class transcription factor and some of these changes are direct effects on transcription as a result of Myc binding to DNA, with or without its binding partner, Max. But other effects are via indirect action. Most notably in Drosophila, in marked contrast to the situation in mammals, Myc stimulation of 18S and 28S rRNA production does not involve enhanced Polymerase I transcription through direct binding of Myc to rRNA gene promoters. Instead, indirect actions of Myc such as enhanced production of Polymerase I co-factors and rRNA processing enzymes stimulate rRNA synthesis. It has been proposed that the Myc-induced changes in ribosome levels and translation rates may increase the protein levels of critical components of the S to G endocycle and thus produce the tight linkage between increased growth and faster DNA endoreplication seen in EP tissues (Zhou, 2020).

Ribosome synthesis is orchestrated in the nucleolus where the rDNA arrays are transcribed and ribosome assembly proceeds within the granular components of the organelle. In Drosophila larval tissues, Myc overexpression greatly increases the size of the nucleolus, but in keeping with its indirect role in rRNA production, Myc protein localizes to many euchromatic sites on the larval salivary gland polytene chromosomes but not to the rDNA arrays. Several of the Drosophila genes identified as being positively transcriptionally regulated by Myc encode nucleolar proteins and two nucleolar components—a DEAD box helicase (Pitchoune) and a protein of unknown function (Nol12/Viriato)–are indicated to act genetically downstream of Myc in EP growth (Zhou, 2020).

The Jim Lovell protein (Lov), previously known as Tkr, is a member of the Tramtrack (Ttk) subset of BTB/POZ proteins in Drosophila. In addition to a single BTB/POZ domain, which acts as a protein interaction interface, most of these proteins contain one or more DNA binding motifs—either of the zinc finger or pipsqueak category. Lov contains a single pipsqueak domain. Like most of the Ttk subgroup, Lov is thus implicated in transcriptional regulation (Zhou, 2020).

The lov gene was initially isolated through its role in gravitaxic behavior in adult Drosophila (Armstrong, 2006) which led to naming it for the heroic Apollo 13 astronaut, Jim Lovell. Subsequently additional behavioral abnormalities were identified through mutant analysis and lov RNAi knockdown. Surprisingly the hypoxic behaviors that were discovered in larvae with tracheal lov knockdown proved to originate from fluid filling of the tracheae due to strong inhibition of tracheal EP growth. This finding led to an investigation of lov function in other endopolyploid larval tissues presented in this study. Lov was found to localize to the nucleolus and regulates EP growth in all the tissues examined. However, epistatis experiments for lov and Myc revealed that lov regulates EP growth differently in different tissues. In the tracheae, lov positively regulates uninflatable, a gene encoding a transmembrane protein of the tracheal cell apical surface which acts downstream of Myc. In contrast, uif plays a has no role in fat body EP growth and the actions of lov in this tissue are all upstream of Myc. These findings uncover heterogeneity in the mechanisms used to orchestrate EP growth throughout the organism (Zhou, 2020).

These studies indicate a role for lov in the growth of all the EP tissues investigated. Myc, like lov, also has a universal role in EP growth but the difference in uif function between the tracheae and the fat body leads to a difference in the relationship of lov and Myc in the two tissues. lov is epistatic to Myc in the tracheae because the action of one of lov’s tracheal targets, is downstream of Myc. But in the fat body, in the absence of a role for uif, loss of lov function has much smaller effect on Myc-induced enhanced EP growth (Zhou, 2020).

Three of the four EP tissues this study investigated (tracheae, salivary glands, epidermis) are epithelial tissues of ectodermal origin. In the two of these tissues (tracheae and salivary glands), lov acts downstream of Myc. Given that the downstream action of lov in one of these tissues involves its target, uif, the possibly was considered that the EP growth of all ectodermally-derived larval tissues is dependent on uif expression regulated by lov. The epidermis is known to express uif in larval life, but uif expression in the salivary gland is limited to the embryonic stage. Other lov targets are therefore likely to act downstream of Myc in the larval salivary gland (Zhou, 2020).

The fat body is derived from the mesoderm, and the cellular effects of lov knockdown in this tissue clearly differ from those in the three ectodermally derived tissues studied. lov plays a lesser role in EP growth with a smaller effect on nuclear size and no detectable effect on cell size. uif knockdown has no effect on fat body EP growth and in contrast to the other tissues, lov acts upstream of Myc in the fat body. It is possible that the action of lov in this tissue involves regulation of tissue-specific genes with very minor roles in EP growth. An alternative explanation involves lov's action at the nucleolus. Lov protein is detected in the nucleoli of multiple EP tissues suggesting that its role at this site is shared between many tissues. The action of lov relative to Myc in the fat body could thus represent the phenotype of lov knockdown when only its nucleolar function is operating in a tissue (Zhou, 2020).

Given that Lov and Myc are both transcription factors one possible underlying mechanism for their interactions would be transcriptional regulation of one gene by the other. The effects were examined of under- or over-expressing each of them on the transcript levels of the other using whole larval RNA preparations, and no evidence was found of cross-regulation at this level. But these data represent a summation of events in all the individual tissues, which could easily obscure tissue-specific responses. A tissue-by-tissue approach will be necessary to fully address this question (Zhou, 2020).

Altogether these studies of the relationship between lov and Myc demonstrate heterogeneity in the responses of individual EP tissues to lov as a regulator of EP growth. As such, they emphasize that EP growth cannot be viewed as a single universal mechanism under the control of the master regulator Myc, but rather as a variable phenomenon, in which tissue-specific growth factors modulate Myc action (Zhou, 2020).

Some insights into how Lov might act at the molecular level can be gained by considering the functions of its close structural relatives. Lov is a member of the Tramtrack (Ttk) BTB/POZ protein subfamily in Drosophila, almost all of whose members also contain at least one DNA binding motif. These proteins are distinctive in that they show unusually high sequence similarity in three regions of the BTB/POZ domain that form an interface for protein-protein binding. This similarity led to the prediction that Ttk proteins generate a network of interacting DNA binding proteins, whose functions require partnering through their BTB/POZ domains. Multiple studies have confirmed this hypothesis. Complexes of Ttk proteins have been shown to regulate both chromatin-related functions and quantitative aspects of certain developmental processes (Zhou, 2020).

These findings thus argue for role(s) for Lov in chromatin structure that involve interaction with other Ttk protein group members. Interestingly Tramtrack has been shown to have a role in the signaling pathway that initiates EP growth in the ovarian follicle cells, where it acts downstream of the cell surface receptor Notch. Tramtrack thus represents one potential binding partner for Lov. No other Ttk family member has been shown to localize primarily to the nucleolus like Lov but Ribbon is known to be required for nucleolar integrity in the larval salivary glands suggesting that it too could be a Lov interaction partner (Zhou, 2020).

Formation of homotypic and heterotypic dimers and oligomers of Lov with other Ttk proteins may also underlie the unexpected discovery that over-expressing lov in various EP tissues produces a more severe version of the lov knockdown phenotype. Over-expressing lov could disrupt its interaction patterns with other Ttk proteins causing loss of its normal activities. Studies to address these possibilities are in progress (Zhou, 2020).

The signals in late embryogenesis that initiate the transition to EP growth in preparation for larval life have not been identified, but for initiation of EP growth in the somatic follicle cells of the adult ovary, the activating sequence has been well-characterized. Upregulation of the Notch ligand Delta in the adjacent germ line cells leads to Notch activation in the follicle cells followed by Notch generated transcriptional events that block the mitotic G2-M transition and promote the G1-S transition of the cell cycle. Myc over-expression is not capable of initiating a switch from mitotic to endocycling in these cells but rather it accelerates the endocycle so that larger cells with larger nuclei are produced. These findings for the role of Myc are consonant with current knowledge of Myc’s role in EP growth in larval cells: it appears to be a downstream activator that coordinates enhanced cellular growth (Zhou, 2020).

In the trachea, the findings indicate that uif acts late in the chain of events that promote EP growth, as a downstream target of the transcription factor Lov, which is itself downstream of the transcription factor Myc. This late function suggests an intracellular role for Uif, but both its structure and cellular location seem at odds with this possibility: Uif is a transmembrane protein of the apical plasma membrane of the tracheal epithelial cells. Its extracellular domain contains a continuous array of 14 EGF-like repeats. This structure evokes the stretches of EGF-like repeats in the extracellular domains of Notch and its ligands Delta and Serrate, suggesting a role for Uif as a membrane signaling molecule that interacts with Notch or another EGF-like repeat containing protein at the cell surface (Zhou, 2020).

Two studies have addressed the significance of the EGF-like repeats of Uif in relation to Notch function. Both found that Uif has no role in the classic plasma membrane signaling function of Notch and currently there is no evidence that the extracellular domain of Uif acts as a receptor to transduce an extracellular signal. However, Loubery (2014) has identified a novel intracellular interaction of Uif with Notch required for the asymmetric distribution of Notch on Sara endosomes during sister socket/sheath cell differentiation. Uif and Notch are internalized independently to these endosomes but their interaction on the endosome surface, through four EGF-like repeats in the Uif external domain, determines the asymmetric inheritance of the Sara endosomes. Intracellular endosome trafficking of Uif has also been demonstrated in the tracheae, with Uif co-localizing internally with Crumbs, another apical transmembrane protein that contains EGF-like repeats (Zhou, 2020).

These intracellular actions of Uif and the known role of Notch in initiating follicle cell EP growth suggest a possible regulatory pathway for activation of tracheal EP growth. A receptor with EGF-like repeats could be the trans-membrane activator, with Uif’s role involving internalization onto a population of endosomes prior to interaction with this protein. Such a site of action for Uif is more consonant with the discovery that uif acts downstream of lov and Myc in the tracheae than a role for Uif in plasma membrane initiation of EP growth (Zhou, 2020).

Failure to burrow and tunnel reveals roles for jim lovell in the growth and endoreplication of the Drosophila larval tracheae

The Drosophila protein Jim Lovell (Lov) is a putative transcription factor of the BTB/POZ (Bric- a-Brac/Tramtrack/Broad/ Pox virus and Zinc finger) domain class that is expressed in many elements of the developing larval nervous system. It has roles in innate behaviors such as larval locomotion and adult courtship. In performing tissue-specific knockdown with the Gal4-UAS system this study identified a new behavioral phenotype for lov: larvae failed to burrow into their food during their growth phase and then failed to tunnel into an agarose substratum during their wandering phase. These phenotypes originate in a previously unrecognized role for lov in the tracheae. By using tracheal-specific Gal4 lines, Lov immunolocalization and a lov enhancer trap line, lov was stablished to be normally expressed in the tracheae from late in embryogenesis through larval life. Using an assay that monitors food burrowing, substrate tunneling and death lov tracheal knockdown was shown to result in tracheal fluid-filling, producing hypoxia that activates the aberrant behaviors and inhibits development. The role of lov in the tracheae that initiates this sequence of events was investigated. When lov levels are reduced, the tracheal cells are smaller, more numerous and show lower levels of endopolyploidization. Together these findings indicate that Lov is necessary for tracheal endoreplicative growth and that its loss in this tissue causes loss of tracheal integrity resulting in chronic hypoxia and abnormal burrowing and tunneling behavior.

The Drosophila BTB domain protein Jim Lovell has roles in multiple larval and adult behaviors

Innate behaviors have their origins in the specification of neural fates during development. Within Drosophila, BTB (Bric-a-brac,Tramtrack, Broad) domain proteins such as Fruitless are known to play key roles in the neural differentiation underlying such responses. Previous work has identified a gene, which was termed jim lovell (lov), encoding a BTB protein with a role in gravity responses. To understand more fully the behavioral roles of this gene this study investigated its function through several approaches. Transcript and protein expression patterns have been examined and behavioral phenotypes of new lov mutations have been characterized. Lov is a nuclear protein, suggesting a role as a transcriptional regulator, as for other BTB proteins. In late embryogenesis, Lov is expressed in many CNS and PNS neurons. An examination of the PNS expression indicates that lov functions in the late specification of several classes of sensory neurons. In particular, only two of the five abdominal lateral chordotonal neurons express Lov, predicting functional variation within this highly similar group. Surprisingly, Lov is also expressed very early in embryogenesis in ways that suggests roles in morphogenetic movements, amnioserosa function and head neurogenesis. The phenotypes of two new lov mutations that delete adjacent non-coding DNA regions are strikingly different suggesting removal of different regulatory elements. In lov⁴⁷, Lov expression is lost in many embryonic neurons including the two lateral chordotonal neurons. lov⁴⁷ mutant larvae show feeding and locomotor defects including spontaneous backward movement. Adult lov⁴⁷ males perform aberrant courtship behavior distinguished by courtship displays that are not directed at the female. lov⁴⁷) adults also show more defective negative gravitaxis than the previously isolated lov^91Y mutant. In contrast, lov⁶⁶ produces largely normal behavior but severe female sterility associated with ectopic lov expression in the ovary. A negative regulatory role is proposed for the DNA deleted in lov⁶⁶ (Bjorum, 2013).

The early Lov staining pattern suggests roles for the protein in three aspects of early embryogenesis. The intense staining in rows of cells that form the boundaries of transient folds in the ectoderm may indicate that Lov functions to promote the mechanical properties of these cells, supporting their role in global re-structuring of the embryo. The strong staining in the dorsal head region, which spans the procephalic neural ectoderm, suggests a role in determination of neural lineages within the head. Finally, the early expression in the amnioserosa could signify a role for Lov in the early differentiation of this extraembryonic tissue and thus its function in dorsal closure and germ band retraction. None of the mutations characterized in this study is a null and none of them affects these early expression elements. Experiments with lov RNAi are therefore being used to address the functions of these early patterns (Bjorum, 2013).

By comparing the Lov expression pattern to that of other early acting genes, this study has identified loci that may be upstream regulators of lov, or act in parallel with it, during these early stages. The three highly-related Dorsocross proteins, which all have the same expression pattern, show a strikingly similar distribution to Lov at stage 8, with the same strong head and amnioserosal staining. This set of proteins does not show the stripe elements seen for Lov at cellular blastoderm but the mRNA expression patterns for pannier (pnr), which encodes a GATA transcription factor and for a gene encoding the serotonin receptor 5HT2 both consist of broad saddles of stripes across the dorsal midline that overlap the Lov saddle along the anterior/posterior axis. Interestingly the 5HT2 receptor has already been shown to play a role in transverse furrow formation in the ectoderm during germ band extension. pnr and the Dorsocross genes are both required for formation of the amniserosa and are regulated by decapentaplegic (dpp) and zernknullt (zen), which act early in the formation of the dorsal embryonic tissues. Both the Zen protein the dpp mRNA pattern also show intense staining in cells at the boundaries of the transient ectodermal folds, as seen with Lov, suggesting that these early acting regulators also control the Lov pattern. The stripe component of the early Lov pattern evokes comparisons to the pair rule and segment polarity genes, whose roles in early pattern formation entail expression in distinct stripes along the anterior-posterior axis . The striped expression of the serotonin receptor 5HT2 is already known to be regulated by the pair rule gene fushi tarazi (ftz) and its ubiquitously expressed cofactor, ftz-F1. Epistatic analyses are planned to determine the position of lov in these early genetic hierarchies (Bjorum, 2013).

The timing and expression pattern of Lov in the PNS indicates that lov acts late in neural development to direct final neuronal differentiation of subsets of neurons. Given the expression pattern of Lov within the various classes of sensory neurons, some elements of the transcriptional hierarchies that regulate lov in the PNS can be predicted. Thus the lov expression in most eso neurons indicates that lov is activated downstream of achaete and scute, the proneural genes of the achaete-scute complex (ASC) that direct formation of the eso mother cells and downstream of cut, which maintains the 'eso' identity in the mother cell lineage. The da class of multiple dendritic neurons, which does not express Lov, also derives from these lineages and so additional regulators must act to prevent lov expression in these neurons. Similarly the absence of Lov in the bp multiple dendritic class suggests suppression of lov by amos, the proneural regulator of this class (Bjorum, 2013).

The expression pattern of Lov within the chordotonal lineages indicates considerable complexity in lov regulation. Limitation of Lov expression to two neurons of the ch class (neurons 2 and 4 of the lateral chordotonal five-neuron cluster) is a striking finding. This is a unique observation that suggests for the first time functional differences amongst the five chordotonal organs (CHOs) at this site. Developmentally, these CHOs are already known to originate via two distinct pathways. Initially, three chordotonal precursors are formed under the direction of the proneural gene atonal. Subsequently rhomboid expression in these precursors leads to EGF receptor signaling, induction of argos in adjacent cells, and formation of two further chordotonal precursors. It is not known precisely which of the final five lateral CHOs correspond to the two derived from the second phase of development but one possibility is that they are the two Lov expressing neurons. Lov expression would thus be downstream of both atonal and EGF signaling in these neurons (Bjorum, 2013).

One of the two CHOs of the ventral cluster (VchA) is also induced by the later wave of EGF signaling activity, but this study found that neither of the two ventral CHOs (VchA and B) expresses Lov, indicating different regulatory mechanisms when compared to lch2 and 4. Further, both the VchA and VchB neurons undergo an additional cell division to generate the two md tracheal neurons td1 and td 2. These neurons, in contrast to their sibling VchA and VchB neurons, express Lov strongly. Generation of these varying Lov expression patterns within the CHO lineages clearly requires a multiplicity of regulatory mechanisms (Bjorum, 2013).

The analyses of lov mRNA expression in the midline lineages performed previously (Kearney, 2004; Stagg, 2011) indicate that regulation of lov within the CNS is also dynamic and complex and that here too lov has roles in the final differentiation of particular neuronal sub-types. The single cell per segment that first expresses Lov protein in the CNS has been identified by the Crews lab as one of the eight primordial midline precursor cells whose determination is controlled by singleminded. Through stages 11–17 of embryogenesis lov mRNA is transiently expressed in the posterior midline glia, the median neuroblast (MNB) cell and a subset of ventral unpaired median motorneurons (VUMs) to give a final expression pattern at stage 17 in just three neurons: a single cell of the MNB progeny and the sister interneuron and motor neuron derived from division of midline precursor neuron 6 (MP6). Notch and lethal-of-scute act at various points to regulate these expression patterns (Bjorum, 2013).

The lov⁴⁷ and lov⁶⁶ mutations produce strikingly different phenotypes, with lov⁴⁷ mutants showing multiple behavioral defects and lov⁶⁶ mutants proving behaviorally unexceptional but strongly female sterile. These differences indicate that the mutually exclusive, non-coding, DNA sequences deleted in the two mutations have differing regulatory roles for the individual lov transcripts. Some of the effects on individual transcripts produced by each mutation generate the differing phenotypes detected in this study but other effects are without consequence, at least in terms of the traits that were assayed. Thus the DNA deleted in lov⁴⁷ has a strong positive role in production of lov transcript D, a lesser positive role in production of transcript A and a negative role in production of transcript C, but the phenotypic effects detected in this study all appear to have their origins in the loss of transcript D alone. Interestingly, transcript B, which has the same transcription start site and expression pattern as transcript D, is not affected by the deletion. It is possible therefore that the embryonic neurons retaining Lov expression in the lov⁴⁷ mutant are neurons that express transcript B as opposed to transcript D (Bjorum, 2013).

It was hypothesized that a global repressor of lov transcript expression in the female gonad is deleted in the lov⁶⁶ mutation and the main phenotypic consequences of this mutation result from loss of this DNA. The deleted DNA also contributes in a positive way to expression of transcripts A, B and D in various tissues and stages. Due to the female sterility, this study did not examine transcript D levels in late embryogenesis for lov⁶⁶ by RT-PCR, but Lov protein expression appeared normal in these stages. However, in adults, lov⁶⁶ depresses neural transcript D expression even more strongly than lov⁴⁷. Given their marked differences in terms of behavioral consequences, it is concluded that the two mutations affect transcript D expression in different subsets of adult neurons. As discussed below, a component of the lov⁶⁶ induced female sterility may result from loss of lov expression in neurons controlling the female genitalia that are unaffected by lov⁴⁷ (Bjorum, 2013).

The larval phenotypes identified for lov⁴⁷ are defects in motor functions: the rates of food shoveling responses and forward locomotion are significantly decreased. The Crews lab has shown that loss of the ventral nerve cord midline neurons, which includes lov expressing cells, results in sluggish larval forward motion, suggesting that this lov⁴⁷ phenotype originates in the CNS. But lov⁴⁷ larvae also show bouts of backward locomotion, which are normally a stimulus-elicited avoidance response. A Central Pattern Generator (CPG) within the CNS coordinates the rhythmic locomotor contraction waves of the larval body wall and prior work has shown that loss of sensory input from the periphery disrupts CPG performance, producing both decreased forward, and spontaneous backward, contraction waves. Evidence has been gathered that most of this peripheral sensory feedback to the CPG derives from the chordotonal neurons of the PNS. Thus, the specific loss of Lov expression in two of the five lateral chordotonal organ neurons in lov⁴⁷ could diminish sensory feedback to the CPG and contribute to abnormal backward movement (Bjorum, 2013).

Motor defects are also seen in lov⁴⁷ adults. The defective lov⁴⁷ gravitactic climb responses cannot be considered a consequence of general sluggishness since lov⁴⁷ adults show enhanced locomotor activity when alone in courtship chambers. They appear to represent a more severe form of the diminished negative gravitaxis produced by the original lov^91Y mutation. Thus the original lov^91Y mutant has normal climb behavior and its decreased negative responses to gravity are only uncovered in the gravitactic maze assay. Like the lov⁴⁷ larval locomotor defects, the lov⁴⁷ defective gravity responses could also reflect loss of lov function in chordotonal neurons. Although the enhancer trap P{GawB} insertion of lov^91Y does not show GAL4 expression in Johnston’s organ, a major graviperceptor organ in the antenna, GAL4 is expressed in the adult leg femoral chordotonal organ, another proprioceptor organ with a demonstrated role in gravity responses in other insects. Thus, the loss of negative gravitaxis in adult lov⁴⁷ mutants could reflect deletion of enhancer sequences adjacent to the lov^91Y P{GawB} insertion point that promote lov expression in the femoral chordotonal organ (Bjorum, 2013).

The larval defects of lov⁴⁷ indicate loss of central organization of locomotor responses. The aberrant courtship responses of lov⁴⁷ males reinforce this concept of failed coordination of innate responses. Male courtship is highly stereotypic with a series of ordered steps (following the female, tapping her, wing vibration, licking the female) leading to abdomen curling and copulation. lov⁴⁷ males showed a marked inability to pursue females continuously but still performed non-directed wing extension, and occasionally abdomen curling, often towards the courtship chamber walls. As noted earlier, Lov belongs to a class of putative transcription factors that includes Fruitless (Fru), the master regulator of male courtship behavior. Given that this class of transcriptional regulators is known to form homo- and hetero-dimers, it is tempting to speculate that Lov might affect courtship responses through protein-protein interaction with Fru. A detailed analysis of Lov protein expression in the adult head has not been undertaken, but preliminary experiments indicate that Lov is expressed in a very large number of adult CNS neurons, a subset of which also express Fru (Bjorum, 2013).

The lov⁶⁶ induced female sterility appears to represent a neomorphic phenotype resulting, at least in part, from ectopic expression of lov in the ovarian germline. However, some aspects of the data for this phenotype are puzzling. First, although lov⁶⁶ hemizygotes show the same phenotype as homozygotes, lov⁶⁶ heterozygotes are essentially wild type both with respect to eggshell features and egg hatch rate. Given that the single copy of the mutant chromosome present in both hemizygotes and heterozygotes should be capable of deregulated lov expression, this result is unexpected. The two genotypes differ however in the state of the lov locus on the homolog paired with the lov⁶⁶ chromosome. In hemizygotes, the entire locus is missing whereas in heterozygotes the presumed regulatory DNA is present on the homolog and could regulate expression from the lov⁶⁶ chromosome via transvection. Recently transvection has been shown to be a common regulatory mechanism in Drosophila, supporting this possibility (Bjorum, 2013).

A further puzzling comparison involves the hatch rates and eggshell defects associated with lov⁶⁶ homozygous/hemizygous mothers and mothers expressing UAS-lov under the ovarian germline GAL4 driver. Whereas for mothers expressing UAS-lov in the germline, the fraction of aberrant eggs (20%) correlates well with the fraction of unhatched eggs (20%), for lov⁶⁶ mothers, the fraction of unhatched eggs (∼80%) is significantly higher than the fraction of detectably abnormal eggs (∼45%). Given that the unhatched eggs from lov⁶⁶ mutant mothers appear undeveloped and unfertilized, one possible explanation is that lov⁶⁶ can disrupt egg maturation and fertilization through a second route, involving other elements of the female reproductive system. Adult females lacking the midline CNS neuron population (which includes lov expressing neurons) show sterility thought to reflect loss of innervation of the female genitalia, perhaps producing failed fertilization. It is possible that lov⁶⁶ causes loss of elements of lov CNS expression in addition to producing ectopic ovarian expression (Bjorum, 2013).

Mechanistically, ectopic expression of lov in the ovary must be presumed to produce interference with the action of one or more factors active in eggshell synthesis and morphogenesis. Making the simplest assumption that a single factor is affected by Lov to produce both phenotypes, this study sought to identify regulators that affect both eggshell synthesis and eggshell morphogenesis and for which action in the follicle cells is controlled by signals from the germline. Two transcriptional regulators, Broad and Tramtrack, meet these criteria. Interestingly these are both BTB transcriptional regulators, offering the possibility, as discussed for Fruitless above, that Lov misregulates function by direct protein interaction. For both Broad and Tramtrack, there is evidence that expression and/or action in the somatic follicle cells is controlled by ecdysone signaling from the germline. Further, mutation of broad and dominant interference with ecdysone signaling produce eggshell phenotypes similar to those seen with lov⁶⁶. In particular 1) the fragile chorions and short/malformed/branched DAs from mothers mutant for the rbp class of broad mutations are similar to the eggs from lov⁶⁶ mothers and 2) the short eggs with stubby DAs and cupshaped dorsal anteriors produced by dominant negative inhibition of ecdysone receptor function in the follicle cells, or loss of ecdysone production, are like the Class 3 eggs identified for lov⁶⁶. These findings suggest that the neomorphic action of Lov in the ovarian germline affects ecdysone signaling and that downstream effects on follicle cell function include disruption of the action of Broad. Although the underlying mechanism of this neomorphic lov⁶⁶ phenotype in detail, this study has determined that, in ovaries from lov⁶⁶ mothers, effects on broad are not limited to effects at the level of protein/protein interaction. broad transcription is depressed in lov⁶⁶ ovaries in addition to disruption of Broad protein localization. Depression of broad expression is consistent with the depressed broad expression seen on loss of ecdysone signaling (Bjorum, 2013).

Search PubMed for articles about Drosophila Jim Lovell

Armstrong, J. D., Texada, M. J., Munjaal, R., Baker, D. A. and Beckingham, K. M. (2006). Gravitaxis in Drosophila melanogaster: a forward genetic screen. Genes Brain Behav 5(3): 222-239. PubMed ID: 16594976

Bjorum, S. M., Simonette, R. A., Alanis, R., Jr., Wang, J. E., Lewis, B. M., Trejo, M. H., Hanson, K. A. and Beckingham, K. M. (2013). The Drosophila BTB domain protein Jim Lovell has roles in multiple larval and adult behaviors. PLoS One 8(4): e61270. PubMed ID: 23620738

Kearney, J.B., Wheeler, S.R., Estes, P., Parente, B., Crews, S.T. (2004). Gene expression profiling of the developing Drosophila CNS midline cells. PubMed ID: 15501232

Loubery, S., Seum, C., Moraleda, A., Daeden, A., Furthauer, M. and Gonzalez-Gaitan, M. (2014). Uninflatable and Notch control the targeting of Sara endosomes during asymmetric division. Curr Biol 24(18): 2142-2148. PubMed ID: 25155514

Pierce, S. B., Yost, C., Britton, J. S., Loo, L. W., Flynn, E. M., Edgar, B. A. and Eisenman, R. N. (2004). dMyc is required for larval growth and endoreplication in Drosophila. Development 131(10): 2317-2327. PubMed ID: 15128666

Stagg, S.B., Guardiola, A.R., Crews, S.T. (2011). Dual role for Drosophila lethal of scute in CNS midline precursor formation and dopaminergic neuron and motoneuron cell fate. Development 138(11): 2171-2183. PubMed ID: 21558367

Zhou, F., Qiang, K. M. and Beckingham, K. M. (2016). Failure to burrow and tunnel reveals roles for jim lovell in the growth and endoreplication of the Drosophila larval tracheae. PLoS One 11: e0160233. PubMed ID: 27494251

Zhou, F., Green, S. R., Tsay, M., Hsu, S., Dibbs, R. and Beckingham, K. M. (2020). The roles of jim lovell and uninflatable in different endopolyploid larval tissues of Drosophila melanogaster. PLoS One 15(8): e0237662. PubMed ID: 32822370

date revised: 12 November 2022

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