As Many Exceptions As Rules: An Immovable Moving Part- That’s Just Cilia!

Biological concepts – primary cilia, sterocilia, kinocilium, Usher syndrome, actin, microtubule, signal transduction, sensory receptor, mechanoreceptor

The USS Oriskany (above) was scuttled in 2006 to

create an artificial reef off of Pensacola Florida. In

2012, the US government effectively ended its policy

of creating artificial reefs this way because of concern

for leaking toxins from the ships to the marine life. But

is was a good way to find a new job for something broken.

Naval vessels are built to move through the oceans. When they can’t, they get fixed or they get decommissioned. As broken vehicles they have no use. Or might they? Some have been re-purposed as man made reefs.

Something that seemed broken because it couldn’t move was given an important new job that didn’t require motility. Remember that analogy as we talk about today’s subject in cilia. Although the order might be reversed.

We spoke last week about how nematodes are the only animals that don’t have cilia. Eukaryotic cilia and flagella (together, the undulipodia) are organelles that move, and in turn may move cells. It turns out that cilia have some exceptions – some don’t beat, and some can’t move at all - so what good are they?

Motile cilia, the kind we have been talking about for the past couple of weeks, are also called 2˚ cilia. If there are 2˚ cilia, I think that pretty much implies that there must 1˚ cilia – and they’re what we will talk about today.

Primary cilia, while less well known, are found on many more cell types than are motile cilia. Motile cilia in mammals are located on male gametes (as flagella), on respiratory epithelium of the lower and upper respiratory tract, fallopian tubes near the ovary and epididymal cells of the testes, and the ependymal cells lining the ventricles of the brain.

Primary cilia are apparent on cells of most types, when they are quiescent (just hanging out, doing it's job). If the cell re-enters the cell cycle and starts to divide or differentiate, the primary cilium will resorb and then reappear in daughter cells once they become quiescent.

Primary cilia have basal bodies and IFT, but their

microtubule structure is different. They don’t have the

inner singlet microtubules, so the outside ones can’t slide

past one another. They do have outer dynein arms, and

those are important for retrograde IFT. See below,

kinocilium don’t even have the outer arms.

Primary cilia are 9(2) + 0 in the axoneme, which they makes them different than motile cilia (see picture and this review). Primary cilia are missing the two central microtubule singlets. They are also missing all dynein, both the inner and the outer arms. This is why they are immotile, sort of.

Another exception with primary cilia is that their microtubule axoneme can change as it goes out to the end of the cilia. It may start out as 9(2) + 0, but at the distal (far) end it's 9 + 0 in nematodes, algae, and in the nose, pancreas and kidneys of vertebrates. All of those count as exceptions too!

In addition, primary cilia are of differing lengths, but most are much shorter than motile cilia. Some don’t even extend from the surface of the cell membrane. However, they're built by IFT (intraflagellar transport) just as 2˚ cilia are, and IFT is important for their functions as well.

So, can a broken cilium have a specific job? If they don’t beat to move a cell or the environment around the cell, then what do primary cilia do? The answer is - just about everything. Primary cilia serve as mechanoreceptors, chemoreceptors, photoreceptors, as well as osmolarity, temperature, or gravity receptors. Think of primary cilia like weather balloons. They stick out into the environment and probe the conditions in the area. They send the data back and the cell can act on it.

As mechanoreceptors, primary cilia might not beat, but they can be moved. They bend in response to flow across the surface and the bend brings a pivot at the level of the basal body – yes, primary cilia have basal bodies just as motile cilia do.

Kidney cells that line the tubules have primary cilia to

sense the urine flow. The cilium is bent, and this triggers

a change in calcium influx. The change is then

transferred to the adjacent cell via calcium channels

that cross both membranes.

The kidney cells that line your urine-filled tubules have primary cilia that stick out into the urine river. As the urine flow speeds up after your third diet coke in the last hour, the primary cilia bend and transmit a signal to the cell. This then signals the cells to ramp up their filtering functions, pulling water back in or excreting urea, etc.

Primary cilia have an asymmetry so that they recognize right from left. In the kidney, the flow is based on orientation, all primary cilia bend in the same direction, toward the anterior. The anterior bend signals for increased calcium influx and then this signal is transmitted to adjacent cells. The uniform gradient (a-p) works cell to cell, and this leads to consistent a-p orientation of the mitotic spindle (which also uses basal bodies in the form of centrioles). The result is that the progeny cells of dividing renal epithelium have the same orientation as the parent they replace.

Back to our nematodes from last week. Primary cilia are the only cilia roundworms have. C. elegans, the roundworm that is used as a laboratory model, is made up of exactly 959 cells – exactly. Sixty of those cells, all sensory neurons, have primary cilia that stick out into the environment via pores called sensillae.

The left photomicrograph has labeled dendrites for sensory

neurons in C. elegans. The right cartoon shows how the

primary cilia from these neurons stick into the pore that

then helps them sense the environment around

the roundworm.

It’s through the interaction of these primary cilia with the worm's immediate environment that it senses its world. This is what passes for a roundworm brain – but your brain has them as well. Especially in the retina of your eyes.

The photoreceptors that absorb light energy and transfer it to electrical impulses are located on a single primary cilium on each retinal cell. The axoneme is used to move photosensitive pigments (like retinal in rhodopsin, see below) back and forth from the receptor to the cytoplasm.

Primary cilia also act as chemoreceptors. In brain proper, they work in formation of new memories – mice without primary cilia can’t remember new objects or recognize objects they have already learned. They can remember the location of the object just fine, just not the object itself. We will talk about primary cilia in the brain much more next week.

Now we can take this discussion a couple of steps further to talk about two ciliary exceptions. There are nonmotile 1˚ cilia, motile 2˚ cilia, and then a third structure called a kinocilium. From the Greek for moving eyelash, the kinocilium is poorly named. Described in guinea pigs in 1989, they don’t move like a blinking eyelash or even like a motile cilium; they lack the inner dynein arms and central microtubules that would allow them to be motile. But, they can move horizontally across a cell surface.

As this movie travels down the photoreceptor, notice the

vertical basal body/axoneme on the left. This is a primary

cilium! The microtubules help move photo pigments up and

down the cilium.

Located in humans on the hair cells the inner ear, kinocilium play a crucial role in both hearing and balance, even though they're gone by the time you hear or need to stand up straight. Their role is regulating the erection of the apparatus that allow hair cells to function.

If hair cell kinocilia are poorly named, then hair cell stereocilia are down right liars. They aren’t cilia at all. The characteristics of cilia include that they are microtubule extensions of a basal body modified from a centriole. They may be motile or nonmotile, but their functions are mediated by moving signaling, structural, or receptor molecules up and down via intraflagellar transport proteins.

None of that applies to sterocilia! They're built from actin not microtubules. They do not have an intraflagellar transport system. They have no basal body. They are very similar to the microvilli of your gut epithelium, but nothing like cilia, except for the fact that they stick up from a cell.

The hair cells work by using the sterocilia as mechanoreceptors. In the cochlea, they bend in response to fluid movement based on vibrations of sound. In the semicircular canals, they bend in response fluid movement as a result of changes in head position. When the sterocilia bend, it generates an action potential in neurons that go to the brain.

Hair cells of the cochlea can be damaged by loud noise.

The left images are the normal (top) hair cell sterocilia, and

the same sterocilia after a loud noise (bottom). The right

images show a series of hair cells in normal condition, and

after a long time exposed to loud noise. Turn down your

music – do you think the hair cells in the damaged cochlea

work well?

So that explains the sterocilia (that aren’t really cilia), but what about the kinocilium? A 2007 paper reviewed how kinocilia mediate production of sterocilia. The hair cells start out with a smooth surface and one long kinocilium in the center of the apical (top) surface. Then the sterocilia start to grow. As the sterocilia appear, the kinocilium moves laterally, to the edge of the apical surface. This defines the orientation of the hair cell – the direction the sterocilia will bend.

The sterocilia start to grow longer, with the ones closest to the kinocilium being the longest. They line up to look like a choir on risers in front of the taller kinocilium. Now they are ready to function. At this point the kinocilium disappears! If you look at working hair cells, you won't find the structure that mediated their development.

So we have two new ciliary structures - neither of which act like cilia. That’s weird enough, but it gets weirder. There is a disease that affects both hearing and vision because it messes with the primary cilia of the retina and the sterocilia of the ear. But we just learned that those are two completely different structures!

People often use "tunnel vision" to explain the field changes

in retinitis pigmentosa, the kind of progressive blindness

in Usher syndrome. But really, it’s more like backing into a

tunnel, one that never reaches the other side. Think of

running this clip backwards.

Called Usher syndrome, victims suffer from hearing loss, vision loss, and balance problems. The vision loss is due to defective maintenance of primary cilia of the retina, but the stereocilia of the cochlea and vestibular system aren’t cilia at all, how could they be affected by a 1˚ cilia protein problem?

As we said last week, nature hates a unitasker. There are at least 11 proteins that work in development and working of both sterocilia AND primary cilia, even though they look and are built completely different. A mutation in one of those proteins affects all three systems. Maybe it would be better if sometimes a protein had just one job, fewer things could get screwed up if something goes wrong with it. Would you rather have reduced vision, reduced hearing, bad balance, or all three?

All this knowledge leaves us with an unanswered question - did the sensory primary cilia develop from motile cilia, or did motile cilia develop from the primary version? Did broken motile cilia develop a new job, or did 1˚ cilia learn how to dance after they had learned their first function? Hmmmm.

We've barely touched the functions of cilia that don’t even move. In the next couple of weeks, we will see how primary cilia keep you from being fat, and how they will be crucial for long-term space travel. Then we can figure out how they give you a right and left hand.

Mathur P, & Yang J (2014). Usher syndrome: Hearing loss, retinal degeneration and associated abnormalities. Biochimica et biophysica acta, 1852 (3), 406-420 PMID: 25481835

Doroquez DB, Berciu C, Anderson JR, Sengupta P, & Nicastro D (2014). A high-resolution morphological and ultrastructural map of anterior sensory cilia and glia in Caenorhabditis elegans. eLife, 3 PMID: 24668170

Patel, A. (2014). The Primary cilium calcium channels and their role in flow sensing Pflügers Archiv - European Journal of Physiology, 467 (1), 157-165 DOI: 10.1007/s00424-014-1516-0

Fry AM, Leaper MJ, & Bayliss R (2014). The primary cilium: guardian of organ development and homeostasis. Organogenesis, 10 (1), 62-8 PMID: 24743231

For more information or classroom activities, see:

Primary cilia –

http://www.nature.com/scitable/topicpage/the-primary-cilium-an-orphan-organelle-finds-14228249

http://jcs.biologists.org/content/123/4/499

http://www.ciliopathyalliance.org/cilia/structure-and-function-of-cilia.html

http://jonlieffmd.com/blog/is-the-primary-cilium-a-cells-antenna-or-its-brain

http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/C/Cilia.html

Hair cells –

http://auditoryneuroscience.com/ear/dancing_hair_cell

http://education-portal.com/academy/lesson/the-ear-sense-of-balance-and-the-inner-ear.html

https://faculty.washington.edu/chudler/hearing.html