Interaction of hookworm 14-3-3 with the forkhead transcription factor DAF-16 requires intact Akt phosphorylation sites

Kiss, Joshua E; Gao, Xin; Krepp, Joseph; Hawdon, John M.

doi:10.1186/1756-3305-2-21

Cited by 14 publications

(26 citation statements)

References 70 publications

(116 reference statements)

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“…Up to 60 minutes of attenuation at 22°C following 60 min at 37°C only caused a slight decrease in expression levels, to 2-fold above the control. As expected, Aca - hsf-1 shows the opposite response as Aca - hsb-1 [21, 43], consistent with the role of Aca -HSB-1 as negative regulator of Aca -HSF-1.…”

Section: Resultssupporting

confidence: 83%

Expression profile of heat shock response factors during hookworm larval activation and parasitic development

Gelmedin

Delaney

Jennelle

et al. 2015

Molecular and Biochemical Parasitology

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When organisms are exposed to an increase in temperature, they undergo a heat shock response (HSR) regulated by the transcription factor heat shock factor 1 (HSF-1). The heat shock response includes the rapid changes in gene expression initiated by binding of HSF-1 to response elements in the promoters of heat shock genes. Heat shock proteins function as molecular chaperones to protect proteins during periods of elevated temperature and other stress. During infection, hookworm infective third stage larvae (L3) undergo a temperature shift from ambient to host temperature. This increased temperature is required for the resumption of feeding and activation of L3, but whether this increase initiates a heat shock response is unknown. To investigate the role of the heat shock in hookworm L3 activation and parasitic development, we identified and characterized the expression profile of several components of the heat shock response in the hookworm Ancylostoma caninum. We cloned DNAs encoding an hsp70 family member (Aca-hsp-1) and an hsp90 family member (Aca-daf-21). Exposure to a heat shock of 42 °C for one hour caused significant up-regulation of both genes, which slowly returned to near baseline levels following one hour attenuation at 22 °C. Neither gene was up-regulated in response to host temperature (37 °C). Conversely, levels of hsf-1 remained unchanged during heat shock, but increased in response to incubation at 37°C. During activation, both hsp-1 and daf-21 are down regulated early, although daf-21 levels increase significantly in non-activated control larvae after 12 hours, and slightly in activated larvae by 24 hours incubation. The heat shock response modulators celastrol and KNK437 were tested for their effects on gene expression during heat shock and activation. Pre-incubation with celastrol, an HSP90 inhibitor that promotes heat shock gene expression, slightly up-regulated expression of both hsp-1 and daf-21 during heat shock. KNK437, an inhibitor of heat shock protein expression, slightly down regulated both genes under similar conditions. Both modulators inhibited activation-associated feeding, but neither had an effect on hsp-1 levels in activated L3 at 16 hours. Both celastrol and KNK437 prevent the up-regulation of daf-21 and hsf-1 seen in non-activated control larvae during activation, and significantly down regulated expression of the HSF-1 negative regulator Aca-hsb-1 in activated larvae. Expression levels of heat shock response factors were examined in developing A. ceylanicum larvae recovered from infected hosts and found to differ significantly from the expression profile of activated L3, suggesting that feeding during in vitro activation is regulated differently than parasitic development. Our results indicate that a classical heat shock response is not induced at host temperature and is suppressed during larval recovery and parasitic development in the host, but a partial heat shock response is induced after extended incubation at host temperature in the absence of a developmental signal, po...

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Section: Resultssupporting

confidence: 83%

Expression profile of heat shock response factors during hookworm larval activation and parasitic development

Gelmedin

Delaney

Jennelle

et al. 2015

Molecular and Biochemical Parasitology

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“…Protein extracts were isolated from activated and non-activated A. caninum L3s as described previously (Kiss et al, 2009). Twelve μg of purified r Ac -HSB-1 were mixed with 100 μg of soluble larval proteins and 485 μL of PBS containing 0.2% Tween 20 (PBS-T) and incubated for 10 h at 4°C on a nutating mixer.…”

Section: Methodsmentioning

confidence: 99%

“…To express Ce -HSF-1 , HEK 293T cells were transfected with 2 μg of pCMV-Tag4/ Ac-hsf-1 plasmid as described (Kiss et al, 2009). After 48 h incubation at 37°C, 5% CO 2 , half of the transfected cells were incubated for an additional 1 h at 42°C while the other half remained at 37°C.…”

Section: Methodsmentioning

confidence: 99%

“…As in C. elegans , hookworm DAF-16 binds to and directs transcription from a conserved DAF-16 binding element, and is regulated by nuclear exclusion following phosphorylation (Gao et al, 2009; Kiss et al, 2009). Given this high level of conservation between their life cycles, we hypothesized that C. elegans and hookworms share a similar HSR pathway.…”

Section: Introductionmentioning

confidence: 99%

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Characterisation of hookworm heat shock factor binding protein (HSB-1) during heat shock and larval activation

Krepp

Gelmedin

Hawdon

2011

International Journal for Parasitology

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When hookworm infective L3s infect their mammalian host, they undergo a temperature shift from that of the ambient environment to that of their endothermic host. Additionally, L3s living in the environment can be exposed to temperature extremes associated with weather fluctuations. The heat shock response (HSR) is a conserved response to heat shock and other stress that involves the expression of protective heat shock proteins (HSPs). The HSR is controlled by heat shock factor 1 (HSF-1), a conserved transcription factor that binds to a heat shock element in the promoter of HSPs, causing their expression. HSF-1 is negatively regulated in part by a HSF binding protein (HSB-1) that binds to and removes HSF-1 trimers bound to HSP gene promoters, resulting in attenuation of the HSR. Herein we describe an HSB-1 ortholog, Ac-HSB-1, from the hookworm Ancylostoma caninum. The Ac-hsb-1 cDNA encodes a 79 amino acid protein that is 71% identical to the Caenorhabditis elegans HSB-1, and is predicted to share the characteristic coiled-coil structural motif comprised of two interacting alpha helices. Recombinant Ac-HSB-1 immunoprecipitated Ce-HSF-1 expressed in mammalian cells that had been heat shocked for 1 h at 42°C, but not from cells incubated at 37°C, indicating that HSB-1 only bound to the active DNA binding form of HSF-1. Expression of Ac-hsb-1 transcripts decreased following 1 h of heat shock, but increased when L3s were incubated at 37°C for 1 h. Activation of hookworm L3s induces an five-to six-fold increase in Ac-hsb-1 expression that peaks at 12 h, coincident with L3 feeding, but that subsequently decreases to two-to three-fold above control at 24 h. Recombinant Ac-HSB-1 immunoprecipitates greater amounts of 70 and 40 kDa proteins from extracts of activated L3s than from non-activated L3s. We propose that an increase in Ac-hsb-1 levels early in activation allows feeding to resume, but that a subsequent decrease in expression permits a HSR that protects nondeveloping L3s at host-like temperatures. Further investigations of the HSR will clarify the role of HSB-1 and HSF-1 in hookworm infection.

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“…Conservation of function was demonstrated by complementation rescue of C. elegans daf-16 mutants (Massey et al, 2006; Hu et al, 2010; Gelmedin et al, 2011) by DNA binding assays (Gao et al, 2009; Gao et al, 2010) and by AKT phosphorylation-dependent binding to the A. caninum 14-3-3 orthologue, FTT-2 (Kiss et al, 2009). The use of purified recombinant Ac-daf-16 DNA binding domain as bait was especially interesting as it identified 24 Ac-daf-16 binding sites, eight of which corresponded to genes differentially expressed on infective larvae activation (Gao et al, 2010).…”

Section: Co-option Of Molecular Pathwaysmentioning

confidence: 99%

The dauer hypothesis and the evolution of parasitism: 20years on and still going strong

Crook

2014

International Journal for Parasitology

116

134

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How any complex trait has evolved is a fascinating question, yet the evolution of parasitism among the nematodes is arguably one of the most arresting. How did free-living nematodes cross that seemingly insurmountable evolutionary chasm between soil dwelling and survival inside another organism? Which of the many finely honed responses to the varied and harsh environments of free-living nematodes provided the material upon which natural selection could act? Although several complementary theories explain this phenomenon, I will focus on the dauer hypothesis. The dauer hypothesis posits that the arrested third-stage dauer larvae of free-living nematodes such as Caenorhabditis elegans are, due to their many physiological similarities with infective third-stage larvae of parasitic nematodes, a pre-adaptation to parasitism. If so, then a logical extension of this hypothesis is that the molecular pathways which control entry into and recovery from dauer formation by free-living nematodes in response to environmental cues have been co-opted to control the processes of infective larval arrest and activation in parasitic nematodes. The molecular machinery that controls dauer entry and exit is present in a wide range of parasitic nematodes. However, the developmental outputs of the different pathways are both conserved and divergent, not only between populations of C. elegans or between C. elegans and parasitic nematodes but also between different species of parasitic nematodes. Thus the picture that emerges is more nuanced than originally predicted and may provide insights into the evolution of such an interesting and complex trait.

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Interaction of hookworm 14-3-3 with the forkhead transcription factor DAF-16 requires intact Akt phosphorylation sites

Cited by 14 publications

References 70 publications

Expression profile of heat shock response factors during hookworm larval activation and parasitic development

Expression profile of heat shock response factors during hookworm larval activation and parasitic development

Characterisation of hookworm heat shock factor binding protein (HSB-1) during heat shock and larval activation

The dauer hypothesis and the evolution of parasitism: 20years on and still going strong

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