Transcytosis

TRANSCYTOSIS: CROSSING CELLULAR BARRIERS

(to appear July, 2003 in Physiological Reviews)

Pamela L. Tuma Ann L. Hubbard

Biology Department Department of Cell Biology

The Catholic University of America Johns Hopkins University School of Medicine

Washington, D. C., USA Baltimore, Maryland, USA

Abstract:

Transcytosis, the vesicular transport of macromolecules from one side of a cell to the other, is a strategy used by multicellular organisms to selectively move material between two environments without altering the unique compositions of those environments. In this review, we summarize our knowledge of the different cell types using transcytosis in vivo, the variety of cargo moved and the diverse pathways for delivering that cargo. We evaluate in vitro models that are currently being used to study transcytosis. Caveolae-mediated transcytosis by endothelial cells that line the microvasculature and carry circulating plasma proteins to the interstitium is explained in more detail, as is clathrin-mediated transcytosis of IgA by epithelial cells of the digestive tract. The molecular basis of vesicle traffic is discussed, with emphasis on the gaps and uncertainties in our understanding of the molecules and mechanisms that regulate transcytosis. In our view there is still much to be learned about this fundamental process.

Table Of Contents:

Introduction
Section I: Documented Transcytosis in vivo
A. Transcytosis in the Vasculature
        1. Structural features of continuous endothelium
        2. Microvascular permeability and transcytosis
B. Transcytosis in the Brain
        1. Cerebral capillaries
                a. Insulin
                b. LDL
                c. Iron
                d. IgG
2. Choroid plexus
C. Immunological Protection and Transcytosis
        1. Structural features of intestinal epithelial cells
        2. M cells, transcytosis and antigen sampling
        3. Transcytosis of IgA
        4. Transcytosis of IgG
D. Role for Transcytosis in the Homeostasis of Micronutrients
        1. Vitamin B12
                a. Uptake from the intestinal lumen
                b. Transfer to circulating TCII and transcytosis
                c. Involvement of additional epithelia in B12 homeostasis
        2. Iron
                a. Transport across the intestine appears not to require vesicular carriers
                b. Uptake of Tf-bound iron
                c. Transcytosis of iron in the placenta and brain
E. Additional Transcytosis Systems
        1. Lung
        2. Mammary gland
        3. Thyroid
F. The Role of Transcytosis in Plasma Membrane Biogenesis in vivo

Section II: In Vitro Cell Models Of Transcytosis
A. What is Constitutes “Good” Transcytotic Cell Model?
B. Microvascular Endothelial Cell Models
C. Epithelial Cell Models
        1. Intestine
        2. Liver
                a. WIF-B cells
                b. Hep-G2 cells
        3. Kidney
        4. Additional Epithelial Cell Systems
                a. Lung
                b. Placenta
                c. Thyroid
                d. Mammary
D. Transcytosis Outside of the Epithelial World
        1. Bone-resorbing Osteoclasts
        2. Neurons

Section III. More About Two Different Transcytosis Systems
A. Caveolae-mediated Transcytosis
        1. The endothelial cell surface
        2. Caveolae and caveolin
        3. Albumin and orosomucoid transcytosis
B. Clathrin-mediated Transcytosis
        1. The pIgA-R transcytotic pathway
        2. Signals and regulation of pIgA-R transcytosis
        3. Transcytosis of pIgA-R versus newly synthesized apical PM residents

Section IV. Mechanisms And Molecules Regulating Transcytosis
A. Targeting Machinery
        1. The SNARE hypothesis
        2. NSF and a-SNAP
        3. t-SNARES and v-SNAREs
        4. Munc18
        5. The rab proteins
        6. The Exocyst
        7. Annexins
        8. Dynamin
B. Cytoskeleton
        1. Microtubules and microtubule-based motors
        2. Actin and actin-based motors
C. Lipids and Transcytosis
        1. PI(3)P
        2. Cholesterol and Glycosphingolipids
D. Perturbations of Transcytosis
        1. Heterotrimeric G Proteins and Protein Kinase A
        2. Calcium, Calmodulin and Protein Kinase C
        3. Possible Mechanisms
E. Transcytosis versus Direct PM Delivery

Conclusion
Acknowledgments
Figure Legends
References
Figures 1-7 and Tables 1-6

Introduction:

At its simplest, transcytosis is the transport of macromolecular cargo from one side of a cell to the other within a membrane-bounded carrier(s). It is a strategy used by multicellular organisms to selectively move material between two different environments while maintaining the distinct compositions of those environments. Cells have other strategies not involving membrane vesicles to selectively move smaller cargo (ions and small solutes) across cellular barriers. Paracellular transport, the movement between adjacent cells, is accomplished by regulation of tight junction permeability, and transcellular transport, the movement of ions and small molecules through a cell, is accomplished by the differential distribution of membrane transporters/carriers on opposite sides of a cell. Together, these three processes contribute to the success of multicellular organisms.

Historically, the existence of transcytosis was first postulated in the 1950’s by Palade iI n his studies of capillary permeability (426) . He described a prominent population of small vesicles, many of which were in continuity with the plasma membrane, and hypothesized that these vesicles were the morphological equivalent of the large pore predicted by the physiologist Pappenheimer to explain the high permeability of blood microvessels to macromolecules (428) . N. Simionescu was the first to coin the term transcytosis to describe the vectorial transfer of macromolecular cargo within the plasmalemmal vesicles from the circulation across capillary endothelial cells to the interstitium of tissues (539) . During this same period, another type of transcytosis was being discovered. Immunologists comparing the different types of immunoglobulins found in various secretions (e.g., serum, milk, saliva and the intestinal lumen), speculated that the form of IgA found in external secretions (called secretory IgA, due to the presence of an additional protein component) was selectively transported across the epithelial cell barrier (577, 578) . The pathway and origin of the component acquired during transport were actively investigated, and in 1980 secretory component (SC) in secretory IgA was identified as the ectoplasmic domain of the intestinal epithelial cell membrane receptor that binds dimeric IgA and transports it through multiple intracellular compartments to the opposite side of the cell (391, 423) . These two historic transcytotic systems are still actively investigated today.

We now know that transcytosis is a wide-spread transport process; a variety of cell types use it, different carriers and mechanisms have evolved to carry it out, and the cargo moved by it is diverse. Cell types: We are most familiar with transcytosis as it is expressed in epithelial tissues, which form cellular barriers between two environments. In this polarized cell type, net movement of material can be in either direction, apical to basolateral or the reverse, depending on the cargo and particular cellular context of the process. However, transcytosis is not restricted to only epithelial cells. Reports of cultured osteoclasts (398, 490) and neurons (221) carrying vesicular cargo between two environments indicate that the strategy of vesicular transcytosis has been used elsewhere. Mechanisms: In intestinal cells transcytosis is a branch of the endocytic pathway, with cargo being internalized via receptor-mediated (i.e., clathrin-coated) mechanisms and progressively sorted away from internalized material destined for other cellular destinations. However, transendothelial transport in blood capillaries does not conform to this scenario, since different carriers and a more direct route are used to cross the cell. Such differences illustrate that multiple transcytotic mechanisms have evolved that depend on the particular cellular context. Further, they illustrate that cargo in the transcytotic pathways seems able to avoid degradation in lysosomes. How? Cargo: The nature of the transcytotic cargo also varies. Although today we might think of transcytosis as a selective process, the originally-defined system, endothelial cells of the microvasculature, moves macromolecular cargo rather non-selectively within the fluid phase of the transport vesicle or by adsorption to the vesicle membrane. Furthermore, transcytotic cargo is not limited to macromolecules. Several vitamins and ions utilize endocytic mechanisms and vesicular carriers as part of their transcellular sojourn. This brings up another unsolved mystery, that of a cell transcytosing particular cargo for use by other cells but also using some of it for its own metabolism. How is such apportionment made?

A major goal of this review is to summarize the widespread occurrence of transcytosis and focus on its many variations. First, we present documented examples of in vivo transcytosis in mammals, using the expanded definition given above. Next, we assess the status of in vitro cell models currently used to study the different types of transcytosis. We then review in more depth the two best-studied transcytosis systems, transendothelial transport of circulating macromolecules and transcytosis of IgA in polarized epithelial cells, focusing on the similarities and differences of their pathways and carriers. Finally, we present current information about the molecular mechanisms and regulation of transcytosis. Throughout, we identify gaps in our present understanding of this process, with the hope that interested researchers will fill in those gaps with insightful experiments and definitive answers.