By David N. Leff
When cell biologists at the University of North Carolina in Chapel Hill set out to study the genetic and antigenic aspects of human cilia lining the bronchial airway tracts, they first studied partial gene sequences - established sequence tags - isolated from a human testis cDNA library. They found that polypeptides encoded by these ESTs showed homology (sequential similarity) to a key ciliary sequence in the genome of sea urchins.
"It turns out that the structural and biochemical composition of cilia are highly conserved in nature," observed cell biologist William Reed, "such that the cilia on a one-celled animal in a pond are very, very similar to the respiratory cilia in human airways. We relied on that high degree of homology in structure and protein sequence to pull out a human relative of the animal gene. It sits on the short arm of chromosome 17."
That far-out sea urchin-testis connection led Reed and his team to demonstrate that this gene is expressed in human airway epithelial cells, which line the respiratory pathways. There, dense arrays of tiny, flexible, hair-like whips beat in concert to sweep mucus-entrapped particles and pathogens up to the throat, which swallows and off-loads this micro-garbage. About 80 percent of the epithelial cells lining healthy human airways are ciliated.
Not that the testis was a far-fetched point of departure. Similar cilia serve to nudge sperm up into the female fallopian tubes, where they meet fertilization-bent eggs pushed down by such cilia from the ovaries.
When airborne pathogens the likes of influenza and pertussis (whooping cough) assail the respiratory tract, their first targets for destruction are those industrious, cleanup-squad cilia. When flu viruses infect the lungs, ciliated cells are damaged and lost. "Normal recovery involves a comeback process that is poorly understood," Reed pointed out
Cilia Are First Casualties Of Microbial Assault
If the pathogen is the Bordetella pertussis bacterium, its target is the trachea, which it strips bare of cleansing cilia. Rapid-fire bursts of coughing try in vain to take over this debris-disposal task. The screeching, spasmodic whoops that follow are gasping to catch the body's breath.
Asthma, a chronic lung disease caused by allergy, not infection, is another victim of ciliary loss. "The repair process can be slowed or halted," Reed noted. "Expression of this gene can be used as a tool to understand the repair process, and how it is interrupted in asthmatics. And we're starting to find that certain molecules that have been associated with asthma prevent the gene from being expressed."
All of the above injured-ciliary ailments - flu, whooping cough, asthma - are reversible. Not so a less common but more deadly hereditary disorder, primary ciliary dyskinesia (PCD). "The cilia of PCD patients," Reed said, "don't move normally. Such people suffer chronic ear and lung infections, and may require lung transplants."
Reed, a post-doctoral fellow, is first author of a paper in the December 2000 issue of the American Journal of Respiratory Cell and Molecular Biology. Its title: "Characterization of an axonemal dynein heavy chain expressed early in airway epithelial ciliogenesis." Reed called this research, completed last year, "groundwork for quantitatively following airway epithelial cell differentiation. This is a good example," he added, "of the way information generated by the Human Genome Project - and 40 years of fundamental cell biology - can be combined and translated into a human research setting. Now that we have located a human gene linked to the appearance of ciliated cells, we have created a new way of following when these cells appear in the lung.
"At the core of each cilium," Reed told BioWorld Today, "is an axoneme - a motile organelle composed of at least 250 polypeptides. Most abundant of these," he went on, "are alpha and beta tubulin, which assemble into nine outer doublet microtubules, and a central pair of singlets." Driving the synchronized beat of the cilia is the axonemal protein dynein, which is powered by ATP - adenosine triphosphate. A dynein consists mainly of one to three heavy chains of protein."
From nine healthy, nonsmoking, adult male volunteers at the university, Reed and his team used bronchoscopy and lung lavage to obtain normal human nasal, tracheal and bronchial epithelium specimens, plus alveolar lung macrophages.
"The probes we used in this study," he pointed out, "have a number of potential applications in addition to the characterization of vertebrate ciliogenesis at a molecular level. They may be useful in the diagnosis and therapeutic treatment of human disorders characterized by congenital ciliary dysfunction, and to isolate ciliated cell-specific gene promoters of value in studying epithelial cell fate during airway maturation, injury and repair or in achieving expression of transgenes."
Recent Work Rounds Out Protein Story
"This journal article," Reed observed, "doesn't report the complete protein sequence our gene expresses. It just shows that the gene is in human airways. Since then," he recounted, "we've gotten the whole protein sequence, so we've been able to determine that. The paper looked at cells in culture, but now we've found that it also turns on in cilia as humans develop. We have followed embryonic tissue at nine, 13, 15 and 18 weeks of gestation in the region that's been known classically as where the airways develop. We were able to show that the gene turns on in those tissues."
Despite his long-term optimism, Reed sees "no immediate prospect of clinical applications. We've obtained such new stuff that we're not sure where it's going to go. It really needs more fundamental work. Studies done in mice we're now doing in humans. So I'm not sure that there are any therapeutic implications - yet."