**31. Knobs and cytoadherence**

A major structural alteration of the host erythrocyte are electron-dense protrusion, or 'knobs', on the erythrocyte membrane of *P. falciparum* cells. The knobs are induced by the parasite and several parasite proteins are associated with the knobs.[75] Two proteins which might participate in knobs formation are the knob-associated histidine rich protein (KAHRP) and erythrocyte membrane protein-2 (*Pf*EMP2), also called MESA. Neither KAHEP nor *Pf*EMP2 are exposed on the outer surface of the erythrocyte, but are localized to the cytoplasmic face of the host membrane. Their exact roles in knob formation are not known, but may involve reorganizing the submembrane cytoskeleton. The knobs are believed to play a role in the sequestration of infected erythrocytes since they are points of contact between the infected erythrocyte and vascular endothelial cells and parasite species which express knobs exhibit the highest levels of sequestration. In addition, disruption of the KAHRP results in loss of knobs and ability to cytoadhere under flow conditions.[76] A polmerophic protein, called *Pf*EMPI, has also been localized to the knobs and is exposed on the host erythrocyte surface. The translocation of *Pf*EMPI to the erythrocyte surface depends in part on another erythrocyte membrane associated protein called *Pf*EMP3.[77] *Pf*EMPI probably functions as a ligand which binds to receptors on host endothelial cells. Other proposed cytoadherence ligand include a modified band-3, called pfalhesin.[78] Seqestrin, rifins and clag9.[79]

*Pf*EMI is a member of the *var* gene family.[80] The 40-50 *var* gene exhibit a high degree of variability, but have a similar overall structure. *Pf*EMPI) has a large intracellular N- terminal domain, a transmembrane region and a C- terminal intracellular domain. The C- terminal region is conserved between members of the *var* family and is believed to anchor *Pf*EMPI to the erythrocyte submembrane cytoskeleton. In particular, this acidic C- terminal domain may interact with the basic KAHRP of the knob[81] as well as spectrin and actin.[82]

The extracellular domain is characterized by 1-5 copies of Duffy-binding (DBL) domains. These DBL domains are similar to the receptor-binding region of the ligand involved in merozoite invasion (discussed above). The DBL domain exhibit a conserved spacing of cystrine and hydrophobic residues, but otherwise show little homology analysis indicates that there are five distinct classes (designated as α,β,γ,δ………… and…ε…..) of DBL domains.[83] The first DBL is always the same type (designated α) and this is followed by a cysteine- rich interdomain region (CIDR). A variable number of DBL in various orders make up the rest of the extracellular domain of *Pf*EMP-I.

#### **Possible Receptors Identified By in Vitro Binding Assays**

• **CD36** 

28 Malaria Parasites

Many proteins that are involved in the invasion process have been identified. However, much still remains to be learned about the cellular and molecular biology of merozoite

Once inside of the erythrocyte, the parasite undergoes a trophic phase followed by replicative phase. During this intra-erythrocytic period, the parasite modifies the host to make it a more suitable habitat. For example, the erythrocyte membrane becomes more permeable to small molecular weight metabolites, presumably reflecting the needs of an

Another modification of the host cell concerns cytoadherence of *P. falciparum*- infected erythrocyte to endothelial cells and the resulting sequestration of the mature parasite in capillaries and post-capillary veinules. This sequestration likely leads to microcirculatory alteration and metabolic dysfunctions which could be responsible for many of the manifestation of severe falciparum malaria. The cytoadherence to endothelial cells confers at least two advantages for the parasite: 1) a microaerophilic environment which is better suited for parasite metabolism, and 2) avoidance of the spleen and subsequent destruction.

A major structural alteration of the host erythrocyte are electron-dense protrusion, or 'knobs', on the erythrocyte membrane of *P. falciparum* cells. The knobs are induced by the parasite and several parasite proteins are associated with the knobs.[75] Two proteins which might participate in knobs formation are the knob-associated histidine rich protein (KAHRP) and erythrocyte membrane protein-2 (*Pf*EMP2), also called MESA. Neither KAHEP nor *Pf*EMP2 are exposed on the outer surface of the erythrocyte, but are localized to the cytoplasmic face of the host membrane. Their exact roles in knob formation are not known, but may involve reorganizing the submembrane cytoskeleton. The knobs are believed to play a role in the sequestration of infected erythrocytes since they are points of contact between the infected erythrocyte and vascular endothelial cells and parasite species which express knobs exhibit the highest levels of sequestration. In addition, disruption of the KAHRP results in loss of knobs and ability to cytoadhere under flow conditions.[76] A polmerophic protein, called *Pf*EMPI, has also been localized to the knobs and is exposed on the host erythrocyte surface. The translocation of *Pf*EMPI to the erythrocyte surface depends in part on another erythrocyte membrane associated protein called *Pf*EMP3.[77] *Pf*EMPI probably functions as a ligand which binds to receptors on host endothelial cells. Other proposed cytoadherence ligand include a modified band-3, called pfalhesin.[78] Seqestrin,

*Pf*EMI is a member of the *var* gene family.[80] The 40-50 *var* gene exhibit a high degree of variability, but have a similar overall structure. *Pf*EMPI) has a large intracellular N- terminal domain, a transmembrane region and a C- terminal intracellular domain. The C- terminal region is conserved between members of the *var* family and is believed to anchor *Pf*EMPI to the erythrocyte submembrane cytoskeleton. In particular, this acidic C- terminal domain

may interact with the basic KAHRP of the knob[81] as well as spectrin and actin.[82]

invasion.[73,74]

**30. Host erythrocyte modification** 

actively growing parasite.

**31. Knobs and cytoadherence** 

rifins and clag9.[79]


Several possible endothelial receptors have been identified by testing the ability of infected erythrocytes to bind in static adherence assays.[84] One of the best characterized among these is CD36, an 88 kDa intergral membrane protein found on monocytes, platelets and endothelial cells infected erythrocytes from most parasite isolates bind to CD36 and the binding domain has been mapped to the CIDR of *Pf*EMPI. However, CD36 has not been detected on endothelial cells of the cerebral blood vessels and parasites from clinical isolates tend to adhere to both CD36 and intracellular adhesion molecule – 1 (ICAMI). ICAMI is a member of the immunoglobulin superfamily and functions in cell-cell adhesion. In addition, sequestration of infected erythrocytes and ICAMI expression has been co-localized in the brain.[85]

Chondroitin sulfate A (CSA) has been implicated in the cytoadhernce within the placenta and may contribute to the adverse effects of *P. falciparum* during pregnancy. The role of some the other potential receptors is not clear. For example, adherence to thrombospondin exhibits a low affinity and cannot support binding under flow condition. Binding to VCAMI, PECAMI and E- selectin appear to be rare and questions about their constrictive expression on endothelial cells have been raised. However, cytoadherence could involve multiple receptor/ligand interactions.

Resetting is another adhesive phenomenon exhibited by *P. falciparum*-infected erythrocytes. Infected erythrocytes from some parasite isolates will bind multiple uninfected erythrocytes and *Pf*EMPI appears to have a role in at least some resetting. Possible receptors include complement receptor-I (CRI), blood group A antigen, or glycosaminoglycan moieties on an unidentified proteoglycan.

Biology of Malaria Parasites 31

and is associated with chromatin modification. This expression spot can only accommodate a single active gene promoter. Thus the var promoter is sufficient for both the silencing and

• The malaria parasite modifies the erythrocyte by exporting proteins into the host cell. • One such modification is the expression of *Pf*EMPI on the erythrocyte surface which

• The binding of this ligand to receptors on host endothelial cells promotes sequestration

• Numerous *Pf*EMPI genes (i.e… the *var* gene family) provide the parasite with a means

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Pollok, JM; *et al*. (2006). "Manipulation of host hepatocytes by the malaria parasite

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• This antigenic variation also correlates with different cytoadherent phenotypes.

the monoclinic transcription of a PfEMPI allele. [96]

and allows the infected erythrocyte to avoid the spleen.

to vary the antigen expressed on the erythrocyte surface.

functions as cytoadherent ligand.

Tindall, ISBN 0-7020-1187-8

*microbiology reviews* 5 (1): 26-35.

**33. Summary** 

**34. References** 

The different types of DBL domains and CIDR (discussed above) bind to different endothelial cell receptors[ 86,87].For example, DBL, which comprises the first domain, bind to many of the receptors associated with resetting. The binding of the CIDR to CD36 may account for the abundance of this particular binding phenotype among parasite isolates.[88]
