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Understanding a severe thymus phenotype in a PKB mouse model: role of specific PKB isoforms in thymus and T cells development

Final Activity Report Summary - IMMUNOKINASE-B (Understanding a severe thymus phenotype in a PKB mouse model: role of specific PKB isoforms in thymus and T cells development)

The immune system enables an organism to resist almost all types of pathogens. It is composed of the closely interacting innate and adaptive immune systems. The innate immune system defends the host in a non-specific manner and provides an immediate defence against pathogens. In contrast, the adaptive immune system allows the specific recognition of a particular pathogen and is responsible for long-term immunity. T lymphocytes are one of the main mediators of the adaptive immune system. Each T cell is characterised by a unique T cell receptor (TCR). TCR allows the T lymphocyte to specifically recognise a given pathogenic antigen. The formation of TCR for each T lymphocyte occurs during T cell development within the thymus, which is therefore essential for the correct generation of a functional T cell repertoire. Hence, failure of normal thymic function eventually results in immunodeficiency.

When hyperactivated, the phosphatidyl-inositol 3 kinase (PI3K) signalling pathway is renowned for its implication in a variety of cancers, justifying the extensive effort placed by pharmaceutical companies to develop therapeutical inhibitors against it. Nevertheless, it is also a critical regulator of numerous physiological processes. Regarding thymocytes (that are T cells developing within the thymus), defects in single components of the PI3K pathway prevent these cells from progressing beyond early T cell developmental stages. The protein kinase PKB is one of the main effectors of PI3K in various physiological and pathological conditions. To determine whether PKB mediates PI3K signalling in the thymus, we characterized this organ in PKB mutant mice.

In mammals, PKB comprises three isoforms: PKBa, PKBb, and PKBg. Of note, at the neonatal age, mice lacking PKBa exhibited a 60 % reduction of thymus weight when normalised to body weight. Hypocellularity, which was not due to increased programmed cell death, accounted for the diminished thymic weight in PKBa-/- neonates and correlated with a decrease in peripheral T cell number. To address whether a block in T cell development could explain thymic hypocellularity in PKBa-/- neonates, we analysed the major thymocyte subsets using flow cytometry. This revealed no striking difference in PKBa-deficient mice compared with age-matched controls. We therefore excluded that a block in T cell development would account for thymic hypocellularity in PKBa-/- neonates. Because reduced thymocyte and peripheral lymphocyte number was specific to the neonatal age, we hypothesized that defective PKB signalling results in developmental delay of the thymus or T lymphocyte progenitors.

A refined analysis in neonatal and adult mice highlighted an accumulation of PKBa-/- thymocytes corresponding to the pre-thymocyte stage. This stage, at which the formation of TCR initiates, is determinant during T cell development. From our data, while PKBa was the main isoform present in pre-thymocyte subsets, PKBb was expressed at significant levels. Of note, when both isoforms were deleted in thymocytes, T cell development was compromised beyond the pre-thymocyte stage. This indicates that PKBa and PKBb play redundant roles during T cell development and suggests that PKB mediates, at least to some extent, PI3K signalling at a key stage of thymocyte development. Extended studies showed that the effect of PKB in thymocytes was cell-autonomous. Moreover, gene expression profile analysis indicated that PKB regulates genes known to respond to (pre)-TCR in pre-thymocytes. These dysregulations likely participate to impaired T cell development.

Thymocytes are a good illustration of the duality of the PI3K-PKB pathway. Our results indicate that signalling mediated by PKB is crucial for the development of thymocytes beyond the pre-thymocyte stage. This stage corresponds to a key checkpoint during T cell development, which is decisive for the generation of a functional T cell repertoire and, therefore, for adaptive immunity. Of note, it has previously been shown that, at the exact same stage, hyperactivation of the PI3K-PKB pathway gives rise to T cell lymphomas. Such a dual role of the PI3K-PKB pathway needs to be taken into account when designing therapeutic drugs to treat diseases such as T cell lymphomas.