RasGRP Ras guanine nucleotide exchange factors in cancer

Abstract

RasGRP proteins are activators of Ras and other related small GTPases by the virtue of functioning as guanine nucleotide exchange factors (GEFs). In vertebrates, four RasGRP family members have been described. RasGRP-1 through -4 share many structural domains but there are also subtle differences between each of the different family members. Whereas SOS RasGEFs are ubiquitously expressed, RasGRP proteins are expressed in distinct patterns, such as in different cells of the hematopoietic system and in the brain. Most studies have concentrated on the role of RasGRP proteins in the development and function of immune cell types because of the predominant RasGRP expression profiles in these cells and the immune phenotypes of mice deficient for Rasgrp genes. However, more recent studies demonstrate that RasGRPs also play an important role in tumorigenesis. Examples are skin- and hematological-cancers but also solid malignancies such as melanoma or prostate cancer. These novel studies bring up many new and unanswered questions related to the molecular mechanism of RasGRP-driven oncogenesis, such as new receptor systems that RasGRP appears to respond to as well as regulatory mechanism for RasGRP expression that appear to be perturbed in these cancers. Here we will review some of the known aspects of RasGRP biology in lymphocytes and will discuss the exciting new notion that RasGRP Ras exchange factors play a role in oncogenesis downstream of various growth factor receptors.

Keywords: Ras; RasGRP; cancer; lymphocytes; receptor; signaling.

Figure 1

Domain structure of RasGRP proteins. REM (Ras exchange motif) and Cdc25 domain form the catalytic core and catalyze GDP to GTP exchange on Ras and Rap GTPases. Two EF hands bind calcium ions and may be important for proper localization and/or GEF regulation. C1 domain of all family members but RasGRP2 binds diacylglycerol and that is crucial for anchoring at the plasma membrane. Finally, RasGRP1 uniquely possess C-terminal tail whose function is unknown but is likely to mediate protein–protein interactions. Conserved phosphorylation sites thought to be important for RasGRPs activation are indicated. All illustrations in this review were made by Anna Hupalowska.

Figure 2

Activation of RasGRP1/3 downstream of TCR/BCR receptor. (A) Overview of TCR-induced RasGRP1-Ras-MAPK cascade. Recognition of the cognate peptide by TCR results in the activation of tyrosine kinase ZAP70, which phosphorylates multiple downstream targets. One of them, the adaptor protein LAT participates in the assembly of a signaling complex containing PLCγ1. PLCγ1 hydrolyses PIP2 present in the plasma membrane into IP and DAG. IP is essential for the release of calcium from internal stores, whereas DAG activates RasGRP1, a GEF for Ras GTPase and initiates MAPK cascade. Note that the composition of the TCR chains and the proximal TCR signaling event are simplified here for clarity. (B) Overview of BCR-induced RasGRP3-Ras-MAPK cascade.Antigen engagement on the BCR leads to activation of the tyrosine kinase Syk. Activated Syk phosphorylates multiple downstream targets, including the adaptor BLNK/SLP-65, which nucleates a signaling complex containing PLCγ2. This leads to hydrolysis of PIP2 into IP3 and DAG. IP3 binds the IP3 receptor on the ER, leading to release of intracellular calcium stores and store-operated calcium entry, while DAG recruits RasGRP1 and RasGRP3 to the plasma membrane and initiates MAPK signaling.

Figure 2

Activation of RasGRP1/3 downstream of TCR/BCR receptor. (A) Overview of TCR-induced RasGRP1-Ras-MAPK cascade. Recognition of the cognate peptide by TCR results in the activation of tyrosine kinase ZAP70, which phosphorylates multiple downstream targets. One of them, the adaptor protein LAT participates in the assembly of a signaling complex containing PLCγ1. PLCγ1 hydrolyses PIP2 present in the plasma membrane into IP and DAG. IP is essential for the release of calcium from internal stores, whereas DAG activates RasGRP1, a GEF for Ras GTPase and initiates MAPK cascade. Note that the composition of the TCR chains and the proximal TCR signaling event are simplified here for clarity. (B) Overview of BCR-induced RasGRP3-Ras-MAPK cascade.Antigen engagement on the BCR leads to activation of the tyrosine kinase Syk. Activated Syk phosphorylates multiple downstream targets, including the adaptor BLNK/SLP-65, which nucleates a signaling complex containing PLCγ2. This leads to hydrolysis of PIP2 into IP3 and DAG. IP3 binds the IP3 receptor on the ER, leading to release of intracellular calcium stores and store-operated calcium entry, while DAG recruits RasGRP1 and RasGRP3 to the plasma membrane and initiates MAPK signaling.

Figure 3

Overview of T cell development and the role of RasGRPs. The expression of CD4 and CD8 marks different stages of T cell development. Early progenitors do not express CD4 or CD8 and are termed double negative (DN). Depending on the expression of other markers those cells are further subdivided into 4 different subsets (DN1–4). DN3 thymocytes express an immature form of the TCR, pre-TCR. Signals from this receptor results in survival, burst of proliferation and differentiation into double positive cells, which express both CD4 and CD8. This process is called β-selection process. Subsequently, signaling from the TCR leads to positive and negative selection, a process depending on avidity of binding to self-peptide-MHC complexes presented by stromal cells in the thymus. Cells differentiate into single positive CD4 or CD8 thymocytes. These cells leave the thymus and migrate to secondary lymphoid organs. RasGRP1 is important for both signaling downstream of pre-TCR as well as for the negative and positive selection. RasGRP4 and 3 also contribute to signaling downstream of preTCR. Note that distinct stages of T cell development are taking places in different anatomical parts of thymus. Also, dynamic patterns of RasGRP1 expression are highlighted-RasGRP1's expression is low in early subsets (DN cells), increases significantly in DP cells and peaks in SP thymocytes to drop again in peripheral T cells.

Figure 4

Overview of B cell development and RasGRP expression. Early B cell progenitors in the bone marrow undergo genetic rearrangement of their immunoglobulin genes, leading to expression of a unique B cell receptor on the surface each B cell. The B cell receptor mediates a series of self antigen-driven checkpoints that progressively eliminate autoreactive clones from the B cell repertoire. As immature B cells progress to a transitional stage, they exit the bone marrow, enter the circulation and migrate to the spleen where their selection and differentiation continues. Splenic B cells differentiate into follicular or marginal zone cells, and mature follicular B cells can recirculate throughout the body and populate the bone marrow and lymph nodes.

Figure 5

Multiple receptor systems couple to different RasGRP proteins. Scheme is showing different receptors which couple to distinct members of RasGRP family. RasGRP1 signals downstream of antigen receptors such as preTCR, TCR, BCR but also from cytokine receptors including G-CSFR and IL7R. More recently it has also been shown that RasGRP1 also couples to Fc binding receptor as well as to GPCR such as CXCR4. RasGRP2 is mainly activated downstream of GPCR such as thrombin receptor. RasGRP3, similarly to RasGRP1 is engaged after antigen receptor triggering, but can also function downstream of receptor tyrosine kinases such as cMet (HGFR) or EGFR and cytokine receptor (GM-SCF). Finally, fMLP receptor (GPCR), FcR, preTCR and GM-SCF can all activated RasGRP4. preTCR-pre T cell receptor; TCR-T cell receptor; BCR-B cell receptor; G-CSFR- granulocyte colony stimulating factor receptor; GM-CSFR- granulocyte-macrophage colony stimulating factor receptor; EGFR- Epidermal growth factor receptor; HGFR- hepatocyte growth factor receptor; GPCR- G protein coupled receptor.

Figure 6

Dysregulated RasGRP1 cooperates with cytokine receptor input in T cell leukemogenesis. T cell leukemia cells expand in the bone marrow in response to growth factors like interleukin 7 (IL7 in green) and take over the space in the cavity (uniform purple cells in the illustration). This expansion leads to a loss of the variety in bone marrow cells, such as blood stem cells (in pink), red blood cells (in red) and fat cells (in yellow) that are normally seen in the bone marrow. The Hartzell et al. study describes how two related, but distinct, genetic alterations in T cell leukemia cells, mutated K-Ras or dysregulated Rasgrp1, both lead to T cell leukemia by responding to IL7 and other signals in different manners.