Ubiquity of Innate Immunity

Determined searches among plant and invertebrate animal phyla for the signature proteins of the highly efficient vertebrate adaptive immune system—antibodies, T-cell receptors, and MHC proteins—have failed to find any homologs. Yet without them multicellular organisms have managed to survive for hundreds of millions of years. The interior spaces of organisms as diverse as the tomato, fruit fly, and sea squirt (an early chordate, without a backbone) do not contain unchecked microbial populations. Careful studies of these and many other representatives of nonvertebrate phyla have found arrays of well-developed processes that carry out innate immune responses. The accumulating evidence leads to the conclusion that multiple immune mechanisms protect all multicellular organisms from microbial infection and exploitation (Table 4-8).

TABLE 4-8 Immunity in multicellular organisms

Taxonomic group	Innate immunity (nonspecific)	Adaptive immunity (specific)	Invasion-induced protective enzymes and enzyme cascades	Phagocytosis	Anti-microbial peptides	Pattern recognition receptors	lympho-cytes	Variable Lympho-cyte receptors	Anti-bodies
Higher plants	+	-	+	-	+	+	-	-	-
Invertebrate animals   Porifera (sponges)   Annelids (earthworms)   Arthropods (insects, crustaceans)	+ + +	- - -	? ? +	+ + +	+ + +	+ + +	- - -	- - -	- - -
Vertebrate animals   Jawless fish (hagfish, lamprey)   Elasmobranchs (cartilaginous fish; e.g., sharks, rays)   Bony fish (e.g., salmon, tuna)   Amphibians   Reptiles   Birds   Mammals	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +	+ + + + + + +

Data from Flajnik, M. F., and L. Du Pasquier. 2008. Evolution of the immune system. In: Fundamental Immunology, 6th ed., W. E. Paul, ed. Lippincott, Philadelphia; and Wong, J. H., L. Xia, Ng, T. B. 2007. A review of defensins of diverse origins. Current Protein and Peptide Science 8:446.

Some Innate Immune System Components Occur across the Plant and Animal Kingdoms

In contrast to the adaptive immune system, components of the innate immune system are evolutionarily ancient, as evidenced by their presence in a virtually all multicellular organisms studied. For example, as mentioned early in this chapter, virtually all plant and animal species, and even some fungi, have antimicrobial peptides similar to defensins. Most multicellular organisms have pattern recognition receptors containing leucine-rich repeats (LRRs), although many organisms also have other families of PRRs. While the innate immune responses activated by these receptors in plants and invertebrates show both similarities and differences compared to those of vertebrates, innate immune response mechanisms are essential for the health and survival of these varied organisms.

Despite plants’ tough outer protective barrier layers, such as bark and cuticle, and the cell walls surrounding each cell, plants can be infected by a wide variety of bacteria, fungi, and viruses, all of which must be combatted by the plant innate immune system. Plants do not have phagocytes or other circulating cells that can be recruited to sites of infection to mount protective responses. Instead, they rely on local innate immune responses for protection against infection. As described in Evolution Box 4-4, some resemble innate responses of animals, while others are quite distinct.

EVOLUTION BOX 4-4

Plant Innate Immune Responses

In the plasma membrane under the cell wall, plant cells express pattern recognition receptors with LRR domains reminiscent of animal TLRs. These PRRs recognize what plant biologists refer to as microbe-associated molecular patterns (MAMPs), including bacterial flagellin, a highly conserved bacterial translation elongation factor, and various bacterial and fungal cell wall components (Figure 1). As is true for animal TLRs, some plant PRRs respond to danger-associated molecular patterns (DAMPs), which usually are created by pathogen enzymes that attack and fragment cell wall components. Some bacterial and fungal pathogens directly inject into plant cells toxin effector proteins that inhibit signaling through the plasma membrane PRRs. These toxins are recognized by a distinct class of LRR receptors in the cytoplasm called R proteins, which, like animal NLR proteins, have both LRR and nucleotide-binding domain (NBD) elements. After ligand binding, plant PRRs activate signaling pathways and transcription factors distinct from those of vertebrate cells (plants do not have NF-κB or IRF homologs), triggering innate responses.

FIGURE 1 Activation of plant innate immune responses. Microbe-associated molecular patterns (MAMPs), danger-associated molecular patterns (DAMPs) generated by microbial enzymes (such as by degrading the cell wall), and microbial effectors (such as toxins) are recognized by plasma membrane LRR-containing pattern recognition receptors (PRRs). Microbial effectors that enter the cytoplasm are recognized by a class of PRRs called resistance (R) proteins. Recognition of MAMPs, DAMPs, and effectors by PRRs induces protective innate immune responses. While there are homologies between plant LRRs and those of animal TLRs, the cytoplasmic domains are very different. For example, some plant PRRs have cytoplasmic domains with tyrosine kinase activity and activate different signaling pathways.

The illustration shows the plant cell wall enveloping the plasma membrane of the plant cell. Several PRRs are on the plasma membrane. The PRRs have a horse-shoe shaped exterior domain protruding into the plant cell wall and an elongated structure extending into the cytoplasm. Some PRRs have a spherical structure attached to the protrusions in the cytoplasm. Bacteria (rod-shaped with filaments), fungi (a cluster of spheres with tubular protrusions on one end), and oomycetes (tubular branches ending in oval nodes) produce MAMPs (represented by circles, triangles, and hexagons) that approach the plant cell wall. DAMPs and cell wall-degrading enzymes produced by bacteria, fungi, and oomycetes intrude into the plant cell wall. The MAMPs, DAMPs, and the cell wall degrading enzymes cause the PRRs to initiate the innate immune responses. The bacteria, fungi, and oomycetes also act as extracellular effectors. The bacteria flagellum penetrates the cell cytoplasm and releases intracellular effectors. Tubular appendages from fungi and oomycetes penetrate the cell wall and reach the plasma membrane, releasing microbial effectors. These intracellular effectors bind to R-protein PRRs that have a horse-shoe shaped exterior domain connected to two spherical structures. Once bound to microbial effectors, the R-protein PRRs initiate innate immune responses.

The primary protective innate immune response mechanisms of plants to infection are the generation of reactive oxygen and nitrogen species, elevation of internal pH, and induction of a variety of antimicrobial peptides (including defensins) and antimicrobial enzymes that can digest the walls of invading fungi (chitinases) or bacteria (b-1,3-glucanase). Plants may also be activated to produce organic molecules, such as phytoalexins, that have antibiotic activity. In some cases, the responses of plants to pathogens even go beyond these protective substances to include structural responses. For example, to limit infection of leaves, PAMP binding to PRRs induces the epidermal guard cells that form the openings (stomata) involved in leaf gas exchange to close, preventing further invasion (Figure 2). Other protective mechanisms include the isolation of cells in the infected area by strengthening the walls of surrounding, noninfected cells and the induced death (necrosis) of cells in the vicinity of the infection to prevent the infection from spreading to the rest of the plant. Mutations that disrupt any of these processes usually result in loss of the plant’s resistance to a variety of pathogens.

FIGURE 2 Induced closure of leaf stomata following exposure to bacterial PAMPs. Under normal light conditions, the openings (stomata) formed by pairs of guard cells in the leaf epidermis are open (visible as the football-shaped openings in the upper pair of leaf photographs), allowing normal gas exchange. However, as shown in the lower panels, exposure of the tobacco leaf to the bacterial plant pathogen Pseudomonas syringae and exposure of the tomato leaf to bacterial LPS induce the closure of the stomata (no openings visible).

The pair of images at the top show the eye-shaped apertures that are the open stomata of the tomato leaf and the tobacco leaf. Two arrows with the text “PAMPs or bacteria" point downward to two images which show the closed stomata of the tomato and the tobacco leaves (eye-shaped apertures cannot be seen).

REFERENCES

• Boller, T., and G. Felix. 2009. A renaissance of elicitors: perception of microbe-associated molecular patterns and danger signals by pattern-recognition receptors. Annual Review of Plant Biology 60:379.

Key Concepts:

• Innate immunity appeared early during the evolution of multicellular organisms.
• Some innate immunity effector functions, such as antimicrobial peptides, are found in invertebrate and vertebrate animals, plants, and even some fungi.
• Innate immune defenses in plants include anatomical barriers as well as PRRs that activate antimicrobial innate immune responses, some of which are similar to those found in animals.

Invertebrate and Vertebrate Innate Immune Responses Show Both Similarities and Differences

Additional innate immune mechanisms evolved in animals, and we vertebrates share a number of innate immunity features with invertebrates (Table 4-8). PRRs (including relatives of Drosophila Toll and vertebrate TLRs) that have specificities for microbial carbohydrate and peptidoglycan PAMPs are found in organisms as primitive as sponges. Together with soluble opsonin proteins (including some related to complement components), some of these early invertebrate PRRs function in promoting phagocytosis. Innate signaling has been well studied in Drosophila, and signaling proteins in flies have been identified that are homologous to several of those downstream of vertebrate TLRs (including MyD88 and IRAK homologs).

One significant difference is that fly Toll does not bind to PAMPs directly. Instead, pathogen binding to soluble pattern recognition proteins activates an enzymatic cascade, the end product of which activates fly Toll. The signaling pathways downstream of Toll are similar to those activated by plasma membrane TLRs of vertebrates. Thus, through the Toll pathway bacterial and fungal infections lead to the degradation of an IkB homolog and activation of NF-κB family members Dif and Dorsal, which induce the production of drosomycin, an insect defensin, and other antimicrobial peptides.

In addition to these and other pathways activated by PRRs, Drosophila and other arthropods employ other innate immune strategies not found in vertebrates, including the activation of phenoloxidase cascades that result in melanization—the deposition of a melanin clot around invading organisms that prevents their spread. Thus invertebrates and vertebrates have common as well as distinct innate immune response mechanisms.

Key Concepts:

• Invertebrates and vertebrates share some innate immune response mechanisms, including phagocytosis and production of anti-microbial proteins and peptides, such as defensins.
• Invertebrate immune responses are best understood in Drosophila melanogaster, where the Toll protein responds to pathogens by activating pathways similar to those downstream of vertebrate TLRs, leading to the production of antimicrobial peptides.
• Invertebrates also use innate immune strategies not found in vertebrates, such as melanization.