Physiological Functions of Mcl-1: Insights From Genetic Mouse Models

Abstract

The ability to regulate the survival and death of a cell is paramount throughout the lifespan of a multicellular organism. Apoptosis, a main physiological form of programmed cell death, is regulated by the Bcl-2 family proteins that are either pro-apoptotic or pro-survival. The in vivo functions of distinct Bcl-2 family members are largely unmasked by genetically engineered murine models. Mcl-1 is one of the two Bcl-2 like pro-survival genes whose germline deletion causes embryonic lethality in mice. Its requisite for the survival of a broad range of cell types has been further unraveled by using conditional and inducible deletion murine model systems in different tissues or cell lineages and at distinct developmental stages. Moreover, genetic mouse cancer models have also demonstrated that Mcl-1 is essential for the survival of multiple tumor types. The MCL-1 locus is commonly amplified across various cancer types in humans. Small molecule inhibitors with high affinity and specificity to human MCL-1 have been developed and explored for the treatment of certain cancers. To facilitate the pre-clinical studies of MCL-1 in cancer and other diseases, transgenic mouse models over-expressing human MCL-1 as well as humanized MCL-1 mouse models have been recently engineered. This review discusses the current advances in understanding the physiological roles of Mcl-1 based on studies using genetic murine models and its critical implications in pathology and treatment of human diseases.

Keywords: Bcl-2; Mcl-1; apoptosis; cell death; genetic mouse model; mitochondria; stem cell.

FIGURE 1

Interactions among Bcl-2 family of proteins confer regulation of the intrinsic apoptotic pathway. Bcl-2 family of proteins are characterized by the presence of Bcl-2 homology (BH) domains. (A) Bcl-2 like pro-survival proteins (Bcl-2, Bcl-x_L, Bcl-w, Mcl-1, and A1/Bfl-1) form a hydrophobic groove that interacts with the BH3 domain of pro-apoptotic proteins. The unique Mcl-1 pro-survival protein is labile and contains a 5’ unstructured region with multiple PEST motifs that are targets for post-translational regulation. (B) The pro-apoptotic proteins can be subgrouped into (i) multidomained proteins (Bak, Bax, and Bok) or (ii) BH3-only proteins (Bid, Bim, Bik, Bad, Bmf, Hrk, Noxa, and Puma). (C) BH3-only proteins are activated upon stress signals. While Bim, Puma, and Bid are able to bind to all pro-survival proteins, Bad binds specifically to and neutralizes Bcl-2, Bcl-x_L, and Bcl-w, and Noxa binds to A1/Bfl-1 and Mcl-1. Bcl-x_L and Mcl-1 preferentially restrain Bak, while all pro-survival proteins are able to bind to Bax. Importantly, all breaks need to be released from Bax and Bak in order for cells to commit to cell death.

FIGURE 2

Established floxed Mcl-1 mouse model systems. (A) Opferman et al. introduced two LoxP sites to the Mcl-1 locus upstream of the ATG start codon, and between exon 1 and exon 2, respectively. (B,C) In the second floxed Mcl-1 model, Bouillet et al. introduced two LoxP sites upstream of the ATG start codon of the Mcl-1 gene, and between exon 3 and 4, respectively. A truncated and non-functional form of hCD4 is introduced downstream of exon 4, which can be subsequently expressed under the control of the endogenous Mcl-1 promoter upon Cre-induced recombination (B). A serendipitous introduction of 13 extra amino acid in the 5’UTR region of the Mcl-1 locus resulted in a more stable form of Mcl-1 protein (C). (D) Dzhagalov et al. introduced LoxP sites flanking exon 1, but the first LoxP site is much further upstream of exon 1 in comparison with the other two floxed Mcl-1 models.

FIGURE 3

Mcl-1 is an essential pro-survival Bcl-2 protein for hematopoietic stem cells (HSC), progenitor cells, and subsets of B lymphocytes. Using multiple conditional knock-out (red solid line) and inducible conditional knock-out systems (blue solid line), Mcl-1 was found to be crucial for the survival of HSC and all the subsets of B cells, including immature and mature B cells, recirculating B, plasma B, and memory B cells.

FIGURE 4

Multiple T lymphocyte lineages and thymic epithelial cells (TEC) rely on Mcl-1 for survival. Conditional knockout (red solid line in the middle panel) and inducible knock-out (blue solid line) models showed that double negative 3 and 4 (DN3 and DN4), double positive (DP), CD4 single positive (SP), CD8 SP, mature T cells, activated T cells, and several subsets of T helper cells are dependent on Mcl-1 for their survival. FoxN1-Cre or Krt5-Cre mediated Mcl-1 deletion in TEC (red solid line in the bottom panel) substantially affected the survival of cortical TEC (cTEC) and medullary TEC (mTEC) and led to a significant decrease of all T cell subsets as the consequence of the disruption of thymic microenvironment (red dashed line).

FIGURE 5

Mcl-1 is indispensable for mature natural killer cells, mast cells, neutrophils, basophils, conventional dendritic cells, and plasmacytoid dendritic cells.

FIGURE 6

Mcl-1 plays an important role for the survival of neurons and neural progenitor cells. In several conditional knock-out models (red solid line), Mcl-1 was shown to be crucial for the survival of neurons in both the developing and post-mitotic neurons in brain.