Signal Transduction

Adaptor proteins are cell signaling molecules linking intracellular proteins, including cell surface receptors to cytosolic effectors. They play key roles in cellular signaling such as phosphorylation, dephosphorylation, signal transduction, organization of the cytoskeleton, cell adhesion, regulation of gene expression, all distinct yet interacting systems. In their pure form, adaptor proteins are devoid of any intrinsic enzymatic activity and serve as intracellular platforms for the amplification and coordinated assembly of multimeric protein complexes. A common feature of adaptor proteins is the organization in modular structures; a limited number of highly evolutionary conserved protein sequences (“domains” or “modules”) are combined to produce a diverse array of protein structures with specific cellular functions and diverse connecting capabilities.

The first phosphoserine/threonine-binding molecules that were identified were members of a family of dimeric proteins called 14-3-3 that were first identified as abundant polypeptides of unknown function in brain; they were later identified as activators of tryptophan and tyrosine hydroxylase and as inhibitors or activators of PKCs. Mammalian cells contain 7 distinct 14-3-3 gene products (denoted β, γ, e, η, σ, t, and ζ), while plants and fungicontain between 2 and 15. Several of the mammalian 14-3-3 isotypes are subject to phosphorylation, although the role that phosphorylation plays in 14-3-3 function remains speculative.

The phosphatidylinositol 3′ –kinase (PI3K)-Akt signaling pathway is an intracellular signaling pathway important in regulating the cell cycle and is activated by many types of cellular stimuli or toxic insults. It regulates fundamental cellular functions such as transcription, translation, proliferation, growth, and survival in response to extracellular signals. This is mediated through serine and/or threonine phosphorylation of a range of downstream substrates.

The Notch signaling pathway is a highly conserved cell signaling system present in most multicellular organisms. Notch signaling plays a pivotal role in the regulation of many fundamental cellular processes such as proliferation, stem cell maintenance and differentiation during embryonic and adult development. The notch cascade consists of notch and notch ligands, as well as intracellular proteins transmitting the notch signal to the cell’s nucleus. In mammalian cells, there are four different notch receptors, referred to as NOTCH1, NOTCH2, NOTCH3, and NOTCH4, which display both redundant and unique functions. Notch receptors are large single pass Type I transmembrane proteins. The extracellular domain of all Notch proteins contains 29–36 tandem epidermal growth factor (EGF)-like repeats, some of which mediate interactions with ligand. After specific ligand binding, the intracellular part of the Notch receptor is cleaved off and translocates to the nucleus, where it binds to the transcription factor RBP-J. In the absence of activated Notch, RBP-J represses Notch target genes by recruiting a corepressor complex. Notch signaling is dysregulated in many cancers, and faulty notch signaling is implicated in many diseases including CADASIL (Cerebral Autosomal Dominant Arteriopathy with Sub-cortical Infarcts and Leukoencephalopathy), T-ALL (T-cell acute lymphoblastic leukemia), Alagille syndrome, MS (Multiple Sclerosis), and myriad other disease states.

Autophagy is an intracellular degradation system that delivers cytoplasmic constituents to the lysosome. Despite its simplicity, recent progress has demonstrated that autophagy plays a wide variety of physiological and pathophysiological roles, which are sometimes complex. Autophagy consists of several sequential steps–sequestration, transport to lysosomes, degradation, and utilization of degradation products–and each step may exert different function. In this review, the process of autophagy is summarized, and the role of autophagy is discussed in a process-based manner.

Receptor Tyrosine Kinases (RTKs) are widely expressed transmembrane proteins that act as receptors for growth factors, neurotrophic factors, and other extracellular signaling molecules. Upon ligand binding, they undergo tyrosine phosphorylation at specific residues in the cytoplasmic tail. This leads to the binding of protein substrates and/or the establishment docking sites for adaptor proteins involved in RTK-mediated signal transduction. RTKs have critical functions in several developmental processes including regulating cell survival, proliferation, and motility. When unregulated, they play prominent roles in cancer formation. The simplest receptor tyrosine kinases (RTKs) have three domains: a ligand binding domain outside the cell, a single membrane-spanning domain, and a tyrosine kinase domain inside the cell. The ligands are usually diffusible peptides or small proteins produced elsewhere in the organism, and are typically growth factors, cytokines and hormones. In the absence of a ligand the receptor is inactive.

Calcium-binding proteins have a vital role in calcium homeostasis by buffering and probably also have a neuroprotective function. Fluctuations in intracellular calcium (Ca2+) are central to orderly neurotransmission and the operation of a wide range of cellular functions. In the lateral BST, a few calbindin D-28k-immunoreactive neurons were scattered in a moderately stained neuropil. Staining of calretinin, another calcium-binding protein, was intense in the rostral pole of the lateral BST, continuous with the caudal ventral striatum. Small calretinin-immunoreactive cell bodies were observed in the lateral BST, but their dendrites could not be clearly visualized because of the intensity of background staining

Cellular senescence, a process that imposes permanent proliferative arrest on cells in response to various stressors, has emerged as a potentially important contributor to aging and age-related disease. A variety of stressors, including strong mitogenic signals, DNA damage, and non-genotoxic chromatin perturbations cause cellular senescence – a state of permanent cell cycle arrest. Cellular senescence, although useful in young organisms to prevent cancer, is thought to promote aging. The most consistent determinant of life-span in eukaryotes is the mitogenic growth hormone/IGF-I pathway. However, premature aging syndromes caused by defects in the cellular response/repair to DNA damage indicate the role of accumulated damage. The complex biology of aging is impacted by both environmental and genetic factors: stochastic DNA damage causes decline of function, and genetics determines the rates of damage accumulation and functional decline.

In humans, circadian rhythms influence the sleep-wake cycle. They also affect other physiological processes such as nervous system activity, hormone production, blood pressure, appetite, digestion and body temperature. Even our mood and cognitive ability is influenced by it. Clock is a basic helix-loop-helix-PAS transcription factor that heterodimerizes with BMAL1. Together these transcription factors bind to E box elements (CACGTG) within the promoter of Period circadian genes (PER1 and PER2) stimulating their transcription. CRY1 is a critical component of the circadian oscillator that represses CLOCK-BMAL1 mediated transcription when translocated into the nucleus. PER1, also known as RIGUI, influences circadian rhythms by interacting with, and stabilizing, other circadian regulatory proteins.

G protein-coupled receptors (GPCRs),G-protein linked receptors, serpentine receptors, or heptahelical receptors are the largest superfamily of membrane proteins in the human genome and they are integral membrane proteins with seven membrane-spanning helices. Upon binding to a ligand – which can range from small molecules like cyclic AMP to peptides and large proteins – GPCRs undergo a conformational change that activates heterotrimeric G proteins (guanine nucleotide-binding proteins), which are important for transmitting the extracellular, ligand signal to the cell interior. G-Proteins can be classified by homologous structure and via common ligand subtypes.

Protein kinases transfer phosphate groups from ATP to serine, threonine, or tyrosine residues on protein peptide substrates, directly affecting the activity and function of the target. Radiolabel studies suggest that approximately 30% of proteins in eukaryotic cells are subject to phosphorylation. Kinase activity, a crucial post-translational modification, regulates a broad range of cellular activities including the cell cycle, differentiation, metabolism, and neuronal communication. In addition, abnormal activity of Akt, ERK, JNK, PKC, PKA, p38, and other MAPK are implicated in many disease states. R&D Systems offers a range of quality products for the detection of protein phosphorylation, which include kinase activity assays, phospho-specific antibodies, ELISA, and more.

ITIMs (immunoreceptor tyrosine-based inhibition motif; S/I/V/LxYxxI/V/L) and ITAMs (immunoreceptor tyrosine-based activation motif; consensus sequence YxxI/Lx6-12YxxI/L) are phosphorylation motifs found in a large number of receptors or adaptor proteins. Phosphorylated ITAMs serve as docking sites for tandem SH2 domains of Syk family kinases, whereas phosphorylated ITIMs recruit tyrosine phosphatases. Signaling through ITAM-bearing receptors usually results in cell activation, while engagement of ITIM-bearing receptors is usually inhibitory, although exceptions have been described. The majority of these receptors are involved in tumor development and regulation of the immune system, although some also function in tissues such as bone and brain.

The JAK/STAT pathway is a principal signaling mechanism for many cytokines and growth factors and provides a direct mechanism to translate extracellular signals into transcriptional responses. Activation of this pathway stimulates cell proliferation, differentiation, migration, growth, survival, apoptosis, and pathogen resistance. These cellular events are critical to hematopoiesis, immune development, growth, adipogenesis, mammary gland development and lactation, and other processes. Mutations that affect JAK/STAT pathway activity are important in inflammatory disease and hematological malignancies, including erythrocytosis and an array of leukemias. JAK inhibitors are also being tested for use in multiple myeloma. Additionally, the JAK/STAT pathway mediates the effects of drug treatments of anemia, thrombocytopenia, and neutropenia, as well as antiviral and antiproliferative agents.

Neurotransmitter receptors are expressed on the surface of post-synaptic cells to bind ligand-specific neurotransmitters and hormones. They are also expressed on presynaptic cells to provide feedback mechanisms and attenuate excessive neurotransmitter release. The majority of neurotransmitter receptors are integral membrane proteins with seven transmembrane domains, commonly coupled to G-proteins. Binding of a ligand to its specific neurotransmitter receptor may result in the activation of a myriad of cell signal transduction pathways and modulation of ion channel homeostasis.

Nuclear factor kappa beta (NFkB) is a nearly ubiquitous pathway responsible for mediating DNA transcription, and therefore cell function. The pathway is activated by a variety of stimuli including cellular stress, cytokines, free radicals, UV radiation, oxidized LDL, and bacterial/viral infection. Activated NFkB is translocated to the nucleus where it binds to specific sequences of DNA called response elements. This DNA/NFkB complex then recruits RNA polymerase and other coactivators, which transcribe downstream DNA into mRNA, for protein synthesis.NFkB1 and NFkB2 are members of the Rel/NFkB family of transcription factors that also includes RelA, c-Rel, and RelB. Rel/NFkB members regulate the expression of genes that participate in immune, apoptotic and oncogenic processes. NFkB is predominantly localized in the cytoplasm as a complex with inhibitory IkB proteins and is released and translocated to the nucleus after phosphorylation of IkB. Both NFkB1 (105 kDa) and NFkB2 (100 kDa) are synthesized as precursor molecules that are proteolytically cleaved to 50 and 52 kDa active subunits. NFkB appears to have contradictory functions in apoptosis and cell survival. NFkB mediates the survival response of many signals by inhibiting p53-dependent apoptosis and up-regulating anti-apoptotic members of the Bcl-2 family, and caspase inhibitors such as XIAP, and FLIP. In contrast, NFkB is also activated by apoptotic stimuli involved in DNA damage and mediates upregulation of pro-apoptotic genes such as TRAIL R2/DR5, Fas, and Fas ligand.

Protein phosphorylation is a reversible and dynamic post-translational modification that is governed by the opposing activities of protein phosphatases and kinases. Protein phosphatases remove phosphate groups covalently attached to serine, threonine, and tyrosine residues. By counteracting the activities of kinases, phosphatases play an important role in the control of a wide variety of cellular functions including cell cycle checkpoints, responsiveness to growth factors, contact inhibition, and cellular motility. Disturbances in phosphatase activity are implicated in a wide variety of disease states such as colon cancer, obesity, and immunodeficiencies. A number of successful drugs have been developed that target protein phosphatases.

Members of the serine proteases are diverse with regard to their function, expression pattern and association with health and disease. Some are active in the coagulation cascade and the complement pathways while others are found specifically in the granules of cytotoxic T lymphocytes and natural killer cells (Granzymes) or in mast cells with trypsin-like specificity (Tryptases). Secreted tissue Kallikreins are cancer markers while transmembrane proteases such as Corin and Enterokinase are convertases. Their proteolytic activity is regulated by different Serpins and receptors such as uPAR and HAIs.