Trib3-deficient mice appear normal under standard

Knockout mice provide valuable information about the functions of a gene at endogenous levels of expression. As such, mice with a targeted disruption of the

Trib3 gene have been generated and studied by several groups (Table 7).

Additionally, several transgenic mouse lines with tissue-specific overexpression of Trib3 have also been created and characterized (Table 8).

Homozygous Trib3 knockout (Trib3−/−_{) mice are viable and physically grossly}

normal, and the crossing of Trib3 heterozygotic (Trib3+/−_{) individuals leads to all}

genotypes being born in the expected Mendelian ratio (Okamoto et al., 2007). These results resemble what has been observed in mice with a deletion of Trib1 or Trib2 (Takasato et al., 2008; Yamamoto et al., 2007). The International Mouse Phenotyping Consortium has recently released phenotyping data from the analysis of adult Trib3 knockout mice, including the characterization of morphology, basic physiology, clinical chemistry, hematology and basic behavioral responses, and currently no significant phenotype associations have been found, although the statistical power may be limited in some cases (data available at: www.mousephenotype.org, accessed 2015-10-29, database release 3.4; Koscielny et al., 2014).

It has been postulated that in the single-knockout mouse lines, other Tribbles homologs might functionally compensate for the lack of one Tribbles family member (Takasato et al., 2008). However, the typical expression patterns of the different Tribbles family members overlap only partially (as discussed in section 2.3.1), and, in Trib3−/− _{mice, unaltered expression of Trib1 and Trib2}

was observed in the liver, white adipose tissue, brown adipose tissue and skeletal muscle (Okamoto et al., 2007). Moreover, functional redundancy would seem more probable between Trib1 and Trib2 than between either of them and

Trib3, since the amino acid sequences of TRIB1 and TRIB2 are considerably

more similar to each other than to TRIB3 (as described in section 2.2.3). Nevertheless, the creation of double- or triple-knockout mouse lines could yield new insights into the functions of the Tribbles homologs in mammals.

Table 7. Characteristics of Trib3-deficient mice.

Phenotype of Trib3 knockout mice Reference

• Viable, physically grossly normal, born in the expected Mendelian ratio

• Unmodified insulin and glucose tolerance

• Normal hepatic insulin signaling, serum biochemistry, body composition and metabolic rate

Okamoto et al. (2007)

• Normal splenic macrophages and lymphocytes Satoh et al. (2013) • On a standard rodent chow diet: unaltered body weight,

blood glucose level and glucose tolerance

• On a high-fat diet: improved glucose tolerance, less body weight gain, lower fasting levels of serum glucose and insulin, decreased liver weight and triglyceride content, unaltered fat pad weight

Koh et al. (2013)

• Unaltered developmental apoptosis of superior cervical

ganglion neurons Zareen et al. (2013)

• Normal baseline kidney function and structure

• With streptozotocin-induced diabetes: increased severity of diabetic kidney disease

Borsting et al. (2014)

• No effect on pancreatic β-cell mass and size

• Normal glucose tolerance, growth and body weight Humphrey et al. (2014) • Low, unaltered baseline incidence of tissue lesions in several

organs at 8 months of age

• In a chemical carcinogenesis model of skin cancer: accelerated malignant progression

• In a genetic model of cancer predisposition: increased incidence of premalignant and malignant lesions in several organs

Salazar et al. (2015)

• No evident alterations in the anatomy or cellular content of the cortical and subcortical regions of the brain

Aime et al. (2015) • Enhanced insulin signaling in brown adipose tissue Jeong et al. (2016) • International Mouse Phenotyping Consortium:

no statistically significant phenotype associations found for adult mice in analyses that span morphology, basic

physiology, clinical chemistry, hematology and basic behavioral experiments (data available at:

www.mousephenotype.org, accessed 2015-10-29) International Mouse Phenotyping Consortium (database release 3.4; Koscielny et al., 2014)

Since previous work by Du et al. (2003) had implicated TRIB3 in insulin signaling in the liver, an organ with a principal role in glucose homeostasis, Okamoto et al. (2007) carried out a thorough characterization of the glucose and energy metabolism of adult Trib3−/− _{mice maintained under standard conditions}

(e.g., feeding with a standard rodent chow diet ad libitum). The results revealed that Trib3-deficient mice have normal serum metabolic parameters and body composition, as well as unaltered hepatic insulin signaling and glucose metabo- lism (Okamoto et al., 2007). Additionally, no discernible effect in insulin or glucose tolerance was found compared to wild type controls, and whole-body metabolic rate and energy expenditure were also not affected by the deletion of

Trib3 (Okamoto et al., 2007). Adequate production of insulin by pancreatic

β-cells is also critical for the control of glucose homeostasis, and an analysis of

Trib3 knockout mice has revealed that they have no apparent differences in

pancreatic β-cell mass and size (Humphrey et al., 2014). Thus, the global loss of

Trib3 does not appear to affect the maintenance of glucose homeostasis in mice

maintained under standard conditions.

Further studies performed with Trib3 knockout mice have revealed that

Trib3−/− _{individuals also have normal baseline kidney function and structure}

(Borsting et al., 2014) and normal macrophage and lymphocyte populations in the spleen (Satoh et al., 2013).

Consistent with the results obtained from Trib3 knockouts, the analysis of transgenic mice that conditionally overexpress human TRIB3 in a Cre recom- binase-dependent manner revealed that ten days of enforced TRIB3 expression in the liver, spleen and kidney causes no apparent histological differences in kidney and spleen (Sakai et al., 2014). However, in the mouse liver, the Cre- mediated overexpression of human TRIB3 gave rise to an increase in hepatocyte nuclear size as well as slight inflammation in the perivascular regions, com- pared to Cre-treated wild type mice (Sakai et al., 2014).