Introduction to the Human Cell. Danton PhD O'Day

Чтение книги онлайн.

СКАЧАТЬ of the components of the sperm has a specific function. Their overall job is to get their payload, the haploid male nucleus, into the egg cytoplasm where it can fuse with the female pronucleus to form the diploid, zygote nucleus.

      Figure 2.9. The structure of the human sperm cell.

      Thus it is clear that each region of the sperm has a specific function. That said, it would also be expected that each of the regions has specific, unique proteins associated with it. However, conjecture in science is of no value without experimentation. So the following experiments were critical to validating these ideas or, if we can be more bold, hypotheses.

      Figure 2.10. Fractions of the human sperm cell were injected into rabbits to make polyclonal antibodies which were then used to stain sperm. This revealed that different proteins were localized in different regions of the sperm cell.

      This work showed:

      •Some proteins are restricted to certain locations

      •Each region (acrosome, nucleus, tail) has specific proteins associated with it

      •These proteins are not found in the other regions (i.e., they are region-specific)

      •It's common for specific regions to serve special functions (e.g. sperm's receptor for egg binding and fusion; junctional adhesion molecules)

      The way specific proteins localize and also form complex protein domains is an exciting area of cell biology that is growing rapidly. For example, RACK1 (Receptor for Activated C Kinase 1) organizes many constituents involved in cellular signaling as covered in a future chapter.

      Lipid Rafts and Caveolae

      Thus proteins are known to exist in domains. But the membrane isn't just made up of a continuous bilayer in which proteins and protein domains reside. There are discontinuities within it. Lipid domains or “rafts” have been shown to exist which contain different concentrations of certain lipids such as cholesterol and sphingolipids. Sphingolipids are a special class of phospholipids involved in cell communication. These rafts are considered to be sites where other specific molecules group for specific functions.

      As indicated in the following figure, caveolae (“little caves”) were first seen in the electron microscope as distinct invaginations (infoldings) of the cell membrane (Figure 2.11). Caveolae are a special type of lipid raft that has small caveolin protein molecules localized on their cytoplasmic side. It is likely that the accumulation of many proteins makes the caveolae lipid rafts detectable in the electron microscope.

      Figure 2.11. Small caveolae which contain the protein caveolin were seen in electron micrographs.

      Caveolae have been implicated in the uptake of cholesterol by endocytosis and in the accumulation of signal transduction and other components prior to their endocytosis by receptor-mediated endocytosis (detailed in Chapter 14). While caveolae are known to be stable, cholesterol-rich membrane domains containing the structure-specific protein caveolin, their potentially diverse roles in cell function are under analysis.

      The membrane components are formed in the Golgi and inserted into the cell membrane but much remains to be learned about their biogenesis. The identification of lipid rafts and caveolae reveals that there is much more to be learned about the structure and function of the cell membrane. Later we’ll discuss how vesicles derived from caveolae can move across epithelial cells in the process of transcytosis, an event that underlies bacterial infections caused by Listeria and other pathogens.

      Fluidity of the Cell Membrane: Early Work

      The “fluid mosaic model” emphasizes the cell membrane is fluid. This “fluidity” wasn't always appreciated by earlier scientists since early models of the cell membrane were very rigid. It took a diversity of early experimentation and observation over many decades to clarify that the cell membrane was indeed fluid. After all, in hind sight, we know that cell division would not be possible with a solid, inflexible membrane. Similarly, skeletal muscle forms by the fusion of cells called myoblasts as covered later in this volume. The sperm and egg must fuse to form a zygote. But fundamental work was required to provide experimental evidence for membrane fluidity. This is summarized in Figure 2.12.

      Figure 2.12. Early experiments revealing the fluid nature of the cell membrane.

      This collection of diverse experiments thus showed:

      •If you cut Amoeba proteus in half with glass needle, both halves survive and crawl away

      •If you stick a glass needle into a frog’s egg, it seals up and the egg remains normal

      •If you treat cells with certain viruses or electricity, cells that don't normally fuse will fuse together

      Membrane Fluidity: Cell Fusion Experiments

      The issue of membrane fluidity took on new meaning when human and mouse cells were induced to fuse and the behavior of their surface proteins was followed (Figure 2.13). The key to understanding this experiment is that antibodies were produced that detected specific proteins unique to either human or mouse cells. These proteins were thus antigens for the specific antibodies. By tagging the antibodies with different fluorescent dyes, it was possible to follow the localization of both the human protein antigen and the mouse antigen. In this experiment the human antibody was coupled with rhodamine, that fluoresces red when viewed under ultraviolet (UV) light with a microscope. The mouse antibody was conjugated to fluorescein isothiocyanate, more commonly called FITC, which fluoresces green under appropriate UV light. These two fluorescent dyes, as well as others, are commonly used in human cell biology research.

      Figure 2.13. The fusion of human and mouse cells showed that proteins in the cell membrane were able to move around.

      When the cells were first fused, the human and mouse antigens remained localized to opposite sides of the fused cell. If the cells were kept cold, this localization changed very little. However, warming the cells to 37^OC led to an intermixing of the fluorescent dyes revealed by a change from separated red and green fluorescence to an intermixed “blue” fluorescence. This verified that the cell membranes were fluid and proteins within them could move around from one area to another.

      In summary, we now know:

      •The lipid phase of the cell membrane, as well as other membranes, is fluid

      •Fluidity depends upon types of lipids, temperature, etc.

      •Membranes fuse during cytokinesis (cell division after mitosis), exocytosis, phagocytosis, etc.

      •Proteins СКАЧАТЬ