Biomedical subjects

Joan E Hooper

Publications and source records attributed to Joan E Hooper.

7 recordsLinked to original sources

PubMed2006

Roadkill attenuates Hedgehog responses through degradation of Cubitus interruptus.

The final step in Hedgehog (Hh) signal transduction is post-translational regulation of the transcription factor, Cubitus interruptus (Ci). Ci resides in the cytoplasm in a latent form, where Hh regulates its processing into a transcriptional repressor or its nuclear access as a transcriptional activator. Levels of latent Ci are controlled by degradation, with different pathways activated in response to different levels of Hh. Here, we describe the roadkill (rdx) gene, which is expressed in response to Hh. The Rdx protein belongs to a conserved family of proteins that serve as substrate adaptors for Cullin3-mediated ubiquitylation. Overexpression of rdx reduced Ci levels and decreased both transcriptional activation and repression mediated by Ci. Loss of rdx allowed excessive accumulation of Ci. rdx manipulation in the eye revealed a novel role for Hh in the organization and survival of pigment and cone cells. These studies identify rdx as a limiting factor in a feedback loop that attenuates Hh responses through reducing levels of Ci. The existence of human orthologs for Rdx raises the possibility that this novel feedback loop also modulates Hh responses in humans.

Alleles↗

PubMed2005

Growth suppression induced by the TRC8 hereditary kidney cancer gene is dependent upon JAB1/CSN5.

TRC8 encodes an E3-ubiquitin ligase disrupted in a family with hereditary renal cell carcinoma (RCC). We previously reported that Drosophila Trc8 (DTrc8) overexpression inhibits growth and that human and fly proteins interact with with the COP9 signalosome (CSN) subunit JAB1/CSN5. However, further mechanistic evidence linking DTrc8 growth suppression to CSN5 was lacking. Here, we show that haploinsufficiency of CSN5, or a T100I point mutation (CSN5(3)), relieved growth suppression by DTrc8, whereas CSN5(1) (E160V) and CSN5(2) (G147D) mutations had no effect. The strength of yeast two-hybrid interactions between DTrc8 and CSN5 were in complete agreement with the observed phenotypes. DTrc8 overexpression resulted in elevated levels of CSN5 and CSN7, but had no effect on NEDD8-modified Cul-1. In contrast to CSN5, heterozygosity for CSN4null had no effect on the DTrc8 phenotype. We also looked for genetic interactions between DTrc8 and other MPN domain proteins in the CSN and 26S proteasome lid. CSN6 haploinsufficiency restored growth, whereas reduction of proteasome subunits RPN8 or RPN11 had no effect. DTrc8 expression increased the level of digitonin-extractable CSN complex, consistent with elevated levels of CSN5 and 7. Our genetic results confirm that DTrc8-induced growth suppression is CSN5 (and CSN6) dependent. While there was no obvious influence on CSN deneddylation activity, the increase in CSN subunits and holocomplex suggests that TRC8 modulates signalosome levels or compartmentalization.

Adaptor Proteins, Signal Transducing↗

PubMed2005

Communicating with Hedgehogs.

Signalling by secreted Hedgehog (Hh) proteins is important for the development of many tissues and organs. Damage to components of the Hh signal-transduction pathway can lead to birth defects and cancer. The Hh proteins are distributed in tissues in a gradient, and cells respond to different thresholds of Hh with distinct responses. The cellular machinery that is responsible for the unique molecular mechanisms of Hh signal transduction has been largely conserved during metazoan evolution.

Animals↗

PubMed2003

Identification of a functional interaction between the transmembrane protein Smoothened and the kinesin-related protein Costal2.

The hedgehog (Hh) family of morphogens plays important instructional roles in the development of numerous metazoan structures. Consistent with the role Hh homologs play in cell fate determination, aberrant Hh signaling results in numerous human pathologies. Hh signal transduction is initiated when Hh binds to its receptor Patched (Ptc), activating the transmembrane protein Smoothened (Smo). Smo transmits its activation signal to a microtubule-associated Hedgehog signaling complex (HSC). At a minimum, the HSC consists of the Kinesin-related protein Costal2 (Cos2), the protein kinase Fused (Fu), and the transcription factor Cubitus interruptus (Ci). In response to HSC activation, the ratio between repressor and activator forms of Ci is altered, determining the expression levels of various Hh target genes. The steps between Smo activation and signaling to the HSC have not been described. Here, we describe a functional interaction between Smo and Cos2, which is necessary for Hh signaling. We propose that this interaction is direct and allows for activation of Ci in response to Hh. This work fills in the last major gap in our understanding of the Hh signal transduction pathway by suggesting that no intermediate signal is required to connect Smo to the HSC.

Animals↗

PubMed2003

Smoothened translates Hedgehog levels into distinct responses.

In the Drosophila wing, Hedgehog is made by cells of the posterior compartment and acts as a morphogen to pattern cells of the anterior compartment. High Hedgehog levels instruct L3/4 intervein fate, whereas lower levels instruct L3 vein fate. Transcriptional responses to Hedgehog are mediated by the balance between repressor and activator forms of Cubitus interruptus, CiR and CiA. Hedgehog regulates this balance through its receptor, Patched, which acts through Smoothened and thence a regulatory complex that includes Fused, Costal, Suppressor of Fused and Cubitus interruptus. It is not known how the Hedgehog signal is relayed from Smoothened to the regulatory complex nor how responses to different levels of Hedgehog are implemented. We have used chimeric and deleted forms of Smoothened to explore the signaling functions of Smoothened. A Frizzled/Smoothened chimera containing the Smo cytoplasmic tail (FFS) can induce the full spectrum of Hedgehog responses but is regulated by Wingless rather than Hedgehog. Smoothened whose cytoplasmic tail is replaced with that of Frizzled (SSF) mimics fused mutants, interfering with high Hedgehog responses but with no effect on low Hedgehog responses. The cytoplasmic tail of Smoothened with no transmembrane or extracellular domains (SmoC) interferes with high Hedgehog responses and allows endogenous Smoothened to constitutively initiate low responses. SmoC mimics costal mutants. Genetic interactions suggest that SSF interferes with high signaling by titrating out Smoothened, whereas SmoC drives constitutive low signaling by titrating out Costal. These data suggest that low and high signaling (1) are qualitatively different, (2) are mediated by distinct configurations of the regulatory complex and (3) are initiated by distinct activities of Smoothened. We present a model where low signaling is initiated when a Costal inhibitory site on the Smoothened cytoplasmic tail shifts the regulatory complex to its low state. High signaling is initiated when cooperating Smoothened cytoplasmic tails activate Costal and Fused, driving the regulatory complex to its high state. Thus, two activities of Smoothened translate different levels of Hedgehog into distinct intracellular responses.

Animals↗

PubMed2002

The TRC8 hereditary kidney cancer gene suppresses growth and functions with VHL in a common pathway.

VHL is part of an SCF related E3-ubiquitin ligase complex with 'gatekeeper' function in renal carcinoma. However, no mutations have been identified in VHL interacting proteins in wild type VHL tumors. We previously reported that the TRC8 gene was interrupted by a t(3;8) translocation in a family with hereditary renal and non-medullary thyroid cancer. TRC8 encodes a multi-membrane spanning protein containing a RING-H2 finger with in vitro ubiquitin ligase activity. We isolated the Drosophila homologue, DTrc8, and studied its function by genetic manipulations and a yeast 2-hybrid screen. Human and Drosophila TRC8 proteins localize to the endoplasmic reticulum. Loss of either DTrc8 or DVhl resulted in an identical ventral midline defect. Direct interaction between DTrc8 and DVhl was confirmed by GST-pulldown and co-immunoprecipitation experiments. CSN-5/JAB1 is a component of the COP9 signalosome, recently shown to regulate SCF function. We found that DTrc8 physically interacts with CSN-5 and that human JAB1 localization is dependent on VHL mutant status. Lastly, overexpression of DTrc8 inhibited growth consistent with its presumed role as a tumor suppressor gene. Thus, VHL, TRC8, and JAB1 appear to be linked both physically and functionally and all three may participate in the development of kidney cancer.

3T3 Cells↗

PubMed1986

Homeotic gene function in the muscles of Drosophila larvae.

The segmental musculature of Drosophila melanogaster larvae consists of 24-30 muscles per segment. Unique patterns of muscles are found in the three thoracic segments and the first and last abdominal segments; the remaining abdominal segments share the same pattern. Mutations in Ultrabithorax (Ubx) cause partial transformation of the muscle pattern of larval abdominal segments towards metathorax. The muscles of the thorax are not affected. In the first two abdominal segments the changes include the loss of at least 11 ;abdominal' muscles and the gain of 11 ;thoracic' muscles. Less extensive transformations are seen in more posterior abdominal segments. Anterobithorax, bithorax, postbithorax and bithoraxoid mutations also induce transformations of the larval musculature. Each allelic group affects a domain that is a subset of the entire Ubx domain but these domains are not restricted to compartments or segments and may extend through as many as five segments. In the muscles the segmental distribution of Ubx antigen correlates with the segments affected by Ubx mutations. The different domains of Ubx in mesoderm and ectoderm argue that the segmental diversity of the muscle pattern is not simply induced by the overlying epidermis and that Ubx function in the mesoderm is required for the correct development of abdominal segments.

Journal Article↗