Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Tribbles

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_135

Synonyms

 NIPK;  SKIP;  TRB;  TRIB

Historical Background

Discovery of Tribbles Genes

Advances in large scale sequencing projects (both cDNA and genome sequencing) combined with the advent of high throughput analytical approaches to detect the targets of certain genes and the molecular consequences of cellular stimulation led to the identification of a number of genes previously unknown to science. A good example of such genes is the tribbles family, where a number of independent strands of research recognized these genes as important regulators and/or targets of cellular function in a range of biological systems.

The name tribbles (TRIB) dates back to a string of papers that published the identification of a novel gene, controlling cell division in early Drosophila development. These reports defined an important regulatory role for tribbles in cell cycle progression during the closure of the ventral furrow (Grosshans and Wieschaus 2000), gastrulation (Seher and Leptin 2000), and oogenesis (Rorth et al. 2000). In these studies, absence of trb allowed the premature mitosis of mesodermal cells thus disrupting gastrulation. Tribbles was also identified as a gene that when overexpressed yielded abnormal numbers of germ cells. These experiments defined a molecular role for trb in cell cycle progression by promoting proteolysis of the Drosophila cdc25 homologues, string and twine which dephosphorylate tyrosine 15 of the mitotic cyclin-dependent kinase cdc2 to promote its activation. Thus, trb coordinates mitosis with cell fate determination, and in the absence of this protein mitosis occurs prematurely. Overexpression of trb in Drosophila caused oocytes to divide more slowly than normal and become abnormally large. Many subsequent research published in mammalian systems is compatible with these early findings and suggests that tribbles proteins represent an important control point of fundamental cellular processes, such as apoptosis, cell division, and cellular activation.

In parallel to the above studies, mammalian tribbles orthologues have been reported, under a variety of names. Tribbles-2 (C5FW) was described as one of the up-regulated genes in dog thyroid in response to mitogens (Wilkin et al. 1996, 1997). Tribbles-1 (C8FW) was then subsequently reported by this group as a close homologue of tribbles-2 (Wilkin et al. 1997). The first study reporting tribbles-3 (Neuronal cell death Inducible Putative Kinase, NIPK) was published by Mayumi-Matsuda et al., demonstrating an increased expression for this gene during neuronal cell death induced by NGF-depletion (Mayumi-Matsuda et al. 1999). These reports were shortly followed by a rapidly increasing body of studies reporting on the functional importance of tribbles, identifying tribbles interacting proteins, and proposing models for their molecular mode of action. This literature has been reviewed systematically in (Docherty and Kiss-Toth 2010; Hegedus et al. 2006, 2007; Kiss-Toth 2005, 2007, 2011).

Below, our current understanding of key aspects of tribbles function will be summarized, as well as some of the key outstanding questions.

Evolution of Tribbles

A comprehensive bioinformatics analysis was performed to identify tribbles orthologues in a range of species and track the origins of this gene family (Hegedus et al. 2006). Systematic analysis of the available EST information in the NCBI database revealed that tribbles-like proteins can already be identified in protozoan species, suggesting that this is an evolutionally ancient, conserved gene. These findings are compatible with the notion that tribbles control fundamental aspects of cellular function. Results of this analysis have also shown that (almost invariably) there is a single tribbles gene per organism in invertebrates. However, all vertebrate species genomes seem to encode for a number of tribbles paralogues. Whilst fish and mammalian species contain three tribbles-like genes, amphibians and birds are missing tribbles-3. Analysis of syntenic regions across vertebrate species suggests that this is due to the loss of a chromosomal segment encoding trb-3 and several other neighboring genes between fish and “modern” amphibians.

Putative Structure of Tribbles Proteins

Tribbles proteins are comprised of three distinct domains (Fig. 1). The N-terminal region of the protein is particularly rich in serine and proline residues, suggesting that this region might be important in protein–protein interactions. It has been shown that this domain is necessary for the nuclear localization of tribbles-1 and -3. The central region of the protein contains an amino acid sequence, which is highly similar to the catalytic domain of serine/threonine kinases. However, some motifs, believed to be crucial to the kinase activity, are missing from tribbles. In particular, kinase sub-domains I, VIB, and VII are absent in tribbles, which suggests that this domain is catalytically inactive and implies evolutionary conservation for structural reasons, necessary for the biological activity of these proteins.
Tribbles, Fig. 1

Alignment of vertebrae tribbles-1/2/3 proteins by using Clustal W method. Red indicates the strongest and black the weakest conservation across the sequences. The N- and C-terminal regions are highlighted in boxes

A single study, using cells under non-stimulated conditions, has addressed the issue of catalytic activity of tribbles, with negative results. However, it remains plausible that tribbles only have kinase activity when activated, as is the case for many serine-threonine kinases.

Intracellular Expression

Most published data on intracellular distribution of tribbles to-date is based on experiments using overexpression systems. Based on these data, tribbles-1 and -3 appear to be localized in the nucleus, whilst tribbles-2 being expressed mostly in the cytoplasm. However, immunostaining of tissues for endogenous tribbles suggests that all three human tribbles orthologues can localize in cytoplasm as well as in the nucleus.

Of note, tribbles expression has been reported to be highly regulated. On one hand, this takes place in a tissue specific manner; thus, expression levels of tribbles family members are tightly controlled during differentiation. Further, cellular stimulation also leads to rapid and often transient changes in tribbles levels. In addition to the control of transcription, tribbles proteins are also thought to be highly unstable, as putative motifs responsible for protein degradation have been predicted in many tribbles orthologues (Hegedus et al. 2007). However, experimental evidence for the stability of tribbles is still somewhat scarce.

Molecular Mechanisms of Tribbles Action

Tribbles Interacting Proteins

A large number of proteins have been reported as tribbles binding partners. These interactions are summarized on Table 1. The relevance of most of these findings is discussed in the appropriate sections below.
Tribbles, Table 1

Summary of tribbles-1/2/3 interacting molecules

Tribbles

Interacting partner

Tissue/cell type

Reference

1

12-lipoxygenase (12-LOX)

Yeast two-hybrid

Tang et al. (2000)

 

MEK-1

HeLa

Kiss-Toth et al. (2004)

 

MKK4

HeLa

Kiss-Toth et al. (2004)

 

C/EBPβ (NK-IL6)

HEK293

Yamamoto et al. (2007)

2

Akt

Liver

Du et al. (2003)

 

C/EBP

U937

Keeshan et al. (2006)

3

Akt

Liver

Du et al. (2003)

 

MEK1

HeLa

Kiss-Toth et al. (2004)

 

MKK7

HeLa

Kiss-Toth et al. (2004)

 

ATF4

Yeast two-hybrid

Bowers et al. (2003)

 

C/EBP homologous protein (CHOP)

HEK293

Ohoka et al. (2005)

 

Bone morphogenic Protein Type II receptor (BMPRII)

Osteoblast, pulmonary aortic smooth muscle cells

Chan et al. (2007)

 

Bone morphogenic regulatory factor 1(Smurf1)

Cos7

Chan et al. (2007)

 

p65/Rel A

HEK293

Wu et al. (2003)

 

PPARγ

HEK293

Takahashi et al. (2008)

 

SIAH1

HEK293T

Zhou et al. (2008)

 

CtIP

HeLa

Xu et al. (2007)

 

Constitutive photomorphogenic protein 1 (COP1)

Adipose tissue

Qi et al. (2006)

 

p300/CREB-binding protein- associated factor (PCAF)

Liver

Yao and Nyomba (2008)

Tribbles Mediated Control of Signal Transduction

Control of signal transduction networks has been suggested as the main mode of action for tribbles. So far, three main concepts have been established.

A number of studies reported that specific pathways of the mitogen activated protein kinase (MAPK) signaling system are controlled by tribbles, much of this specificity is thought to be due to the tissue specific differences in tribbles expression and/or the altered expression of tribbles binding partners. In general, tribbles proteins are thought to bind to the middle layer of the three MAPK cascades and thus control the activity of MAPKK proteins (Eder et al. 2008; Gilby et al. 2010; Sung et al. 2007). These interactions and their biological consequences have mainly been investigated in the context of regulating inflammatory responses of vascular and immune cells.

The second major concept by which tribbles are proposed to regulate cell function is via their interaction with Akt/PKB serine-threonine kinases. It has been suggested that the insulin activated PI3K/PKD/Akt signaling pathway is controlled by this interaction, namely, by a polymorphic variant (Q84R) of tribbles-3 that can affect Akt binding. However, the molecular data on the functional importance of this interaction are somewhat conflicting, some studies demonstrating that trb-3 inhibits Akt activity, whilst others found no modulation of Akt-dependent pathways. It is likely that these discrepancies have arisen due to differences in the cell types and/or stimuli used in the various studies, which would also reinforce the notion that intracellular signals are transmitted via large multi-protein complexes and the exact composition of these may be cell type specific. Thus, trb-3/Akt interaction may have different functional consequences, depending on the microenvironment.

Finally, tribbles are thought to control the turnover of other proteins via their interaction with specific ubiquitin ligases. The first line of evidence for such a regulatory mechanism came from Drosophila studies, published by Roth et al., demonstrating that the turnover of fly C/EBP (slbo) protein is controlled by the Ubp64 ubiquitin ligase, via a tribbles-dependent mechanism (Rorth et al. 2000). Recently, a similar model was put forward in a mammalian setting, reporting that tribbles-1 and -2 but not tribbles-3 are able to promote degradation of C/EBP (Dedhia et al. 2010).

Tribbles Mediated Control of Gene Expression

In addition to interacting with signaling proteins, tribbles have also been reported to bind to several transcription factors and control their transcriptional activity. The first and so far best characterized example of such studies was published by Ord, where they showed that tribbles-3 binds to ATF4 and inhibits the transcriptional activity of this protein (Ord and Ord 2003). Interestingly, whilst tribbles -1 and -2 appear to promote degradation of C/EBP proteins, and tribbles-3 controls ATF4, these two transcription factors seem to cooperate and control the expression of tribbles-3 itself (Ohoka et al. 2005). Putting these findings together, it is likely that complex regulatory feedback loops exist whereby members of the tribbles protein family regulate each other’s expression.

Tribbles and Disease

Since their discovery, tribbles have been associated with a number of human diseases, ranging from diabetes, lipid disorders, cardiovascular disease, cancer as well as neurological disorders, such as narcolepsy. Many of these observations were initially correlative and have later been backed-up by functional studies, as summarized below.

Type 2 diabetes: Shortly after the initial characterization of Drosophila tribbles, a paper published by Marc Montminy’s group have shown that tribbles-3 interacts with unphosphorylated Akt and reduces its activity (Du et al. 2003). Thereby, tribbles-3 was suggested as a novel, stress induced negative regulator of insulin signaling. A follow-on work from this group reported that trb-3 levels were reduced in liver via PPARα and that this is a mechanism to improve glucose tolerance, thus maintain glucose homeostasis. In line with these studies, a polymorphic variant (Q84R) of human tribbles-3 was subsequently identified as a risk factor for the development of type 2 diabetes (Prudente et al. 2009). However, there are also published reports, which suggest that the physiological action of tribbles-3 might be more complex than suggested by these models. A study using primary rat hepatocytes found no role for tribbles-3 in insulin-mediated control of glucose homeostasis (Iynedjian 2005). Further, tribbles-3 deficient mice appear to have a normal glucose tolerance and insulin signaling (Okamoto et al. 2007). Therefore, whist there is strong evidence that tribbles-3 might play a role in the control of insulin signaling and thus contribute to the development of type 2 diabetes, its action is likely to be influenced by other, yet undefined factors.

Lipid metabolism, hyperlipidemia: A molecular study of spontaneous mutations in mice revealed a disorder called fatty liver dystrophy, which is characterized by transient hypertriglyceridemia and fatty liver during the neonatal period, followed by development of a peripheral neuropathy. Dysregulation of tribbles-3 (fld2) has been identified in this mouse strain (Klingenspor et al. 1999), which raised the possibility that elevated trb-3 levels may contribute to the development of this syndrome. However, there is a clear need for further studies to explore the validity of this hypothesis.

Recent genome-wide association studies (GWAS) have identified a number of genes, not previously associated with dysregulated lipid levels in humans. One of these novel candidates is tribbles-1, which was first associated with serum triglyceride levels (Kathiresan et al. 2008) and subsequently with elevated total- and LDL-cholesterol, as well as with an increased risk of coronary heart disease (Tai et al. 2009).

Cancer: An increasing body of evidence has accumulated in recent years to suggest an important role for tribbles in the development of cancer. This is not entirely surprising, given the original observations in Drosophila, where tribbles was shown to inhibit cell cycle progression. However, the role tribbles play in the mammalian setting is still somewhat controversial, as discussed below.

A number of cancer types (including colon cancer, acute myeloid leukemia (AML), gliomas, mammary tumors, etc.) have been reported, in which the expression of tribbles proteins appears to be dysregulated. However, the best-studied example is one of AML. Nakamura and his team have suggested that tribbles-1 cooperates with Hoxa and Meis1 in promoting AML. This group as well as work published by Warren Pear and his colleagues suggested a similar model for tribbles-2 and proposed the tribbles-dependent degradation of C/EBP as the molecular mechanism for this. However, a recent analysis of a large microarray dataset over 300 of human AML samples revealed that tribbles-1 and -2 are expressed at particularly low levels in groups where C/EBP levels were reduced. Thus, further work is clearly needed to address the molecular contribution of tribbles in the development of AML.

Narcolepsy: The latest in the list of diseases associated with tribbles is a chronic sleep disorder, called narcolepsy. It has been long suggested that autoimmunity may contribute to the onset of this disease but thus far no autoantigens were identified. However, recent reports from three independent groups have identified that the presence of anti-tribbles-2 antibodies is associated with this disease. However, the mechanism by which this contributes to disease development is currently unclear.

Summary

Tribbles proteins have become of substantial interest for biomedical research over the last decade, as they appear to control the action of key signal transduction pathways, regulate the turnover as well as the action of transcription factors. As they are believed to be pseudokinases, tribbles are most probably exerting their function via interacting with other proteins and modulating their activity. Whist there is a rapidly increasing body of literature to underline their importance in the development of human disease, many basic aspects of their action and the mechanism by which they contribute to the normal physiology and pathology of these diseases is currently unclear and will require further research.

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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Department of Cardiovascular ScienceUniversity of SheffieldSheffieldUK