Embryology   Biology 441   Vertebrate Embryology, Spring 2014   Albert Harris

 

Monday March 17 Lecture: Induction

Continuing the concept of "Induction", in the special meaning of embryonic induction, which is stimulation of a change or difference in cell differentiation caused by some signal from other cells.

Warren Lewis (~1908) discovered that transplanted (amphibian) embryonic eye cups induce skin ectoderm to differentiate into a normal lens, with the same differentiated cell types and geometric arrangement and shape as the lens of a normal eye.

Hilde Prösholdt and Hans Spemann (1922) discovered that transplanted salamander dorsal lip of the blastopore (cells that will differentiate (and physically rearrange) to form the notochord), if transplanted under the ectoderm in any other part of the embryo will induce formation of all three germ layers to develop into a whole additional body. The notochord induces somatic ectoderm to switch fate and become neural tube ectoderm. Notochord cells also induce mesoderm cells to become somites, and induce endoderm to form digestive tract. Research focused mostly on induction of neural tube by notochord, with less attention to notochord's effects on other germ layers. The dorsal lip of the blastopore is often called "Spemann's Organizer", and this particular example of induction has often been called "primary induction"; but I promise not to ask exam questions requiring these particular names. Concepts, example and significance are the important things.

Induction wouldn't have deserved a Nobel Prize if it were just a method for causing freak salamanders. The importance of induction is that it is the normal mechanism by which the neural tube, somites, etc. are caused to differentiate at their correct geometric locations relative to each other. Likewise, the example of induction earlier discovered by Warren Lewis is the normal mechanism that causes lenses to differentiate from what would otherwise become skin epithelium (and causes neural retina to differentiate from tissue that would otherwise differentiate into pigmented retina epithelium).

In the mouth, odontoblasts (that make dentine) induce differentiation of ameloblasts (that make enamel). If you remove either, then the other doesn't differentiate. If you put an impermeable barrier between the precursors of these two cell types, neither will differentiate without the other. If a mutation were to prevent production of whatever signal molecule odontoblasts secrete that induces formation of ameloblasts, then neither cell type will form, and that animal will have no teeth (even if the inductive signal is still being sent).

Indirect, but very dramatic, evidence seems to prove that lack of that particular inductive signal is how modern birds evolved toothlessness. (In the age of dinosaurs many/all? species of birds had teeth.) The evidence is that chicken embryos will form teeth if you graft mouse odontoblasts into their mouths. Apparently birds retained almost all the genes needed to make teeth, including even the ameloblasts' receptors for whatever signals get sent to them from precursors of odontoblasts. Visualize all those receptor molecules, plus all those genes for making enamel, waiting in the mouths of one bird after another for about seventy million years. I wonder if snake mesoderm is waiting for the right signal to form legs.

More than three hundred photographs and diagrams of induction are on the Google Images web site on the topic "Spemann's Organizer".

Google Images, Spemann's Organizer

Spemann won a Nobel Prize (1935) for this discovery. Dozens (hundreds?) of other examples of embryonic induction have been discovered since Lewis, Prösholdt and Spemann discovered the general phenomenon of cell to cell signals that switch embryonic cells from one differentiated cell type to another.

Several entire issues of research journals have been published as memorials to this great turning point in the history of biology. A good and freely accessible journal volume about induction is Volume number 45, number one of the International Journal of Developmental Biology.

Incidentally, each of their special issues starts with biographical articles about various breakthrough discoveries, and what the researchers were like.

Every case in which cell differentiation depends on any message received from very close cells, embryologists call that a case of induction. Broad use of this word should not be interpreted as a claim that the mechanism of all cases of induction is the same, and especially not that the signal molecules are always the same. They often are the same, however.

Researchers had expected/hoped to develop a bioassay to identify which specific notochord protein signals to ecdoderm to become neural, instead of skin. (Skin was assumed to be the default differentiation; but now there is evidence that nerves are.)

People hoped for something simpler, like auxin, one specific stable chemical from notochord that would turn skin into nerves, turn lateral plate mesoderm into somites, etc.

A big problem was non-specificity. (salami, cell breakdown products, etc.) Other problems were low concentrations and instability of the real signal proteins, like noggin.

Most of these problems were eventually solved by making c-DNAs from embryo tissues (That is, by isolate RNA from tissues that emit inductive signals, then use reverse transcriptase to synthesize the equivalent of messenger RNA. Chordin, Noggin, Follistatin, were discovered that way.)

BMP (Bone Morphogenetic Protein, which had already been discovered during research unrelated to induction) was discovered to stimulate ectoderm to differentiate into skin rather than nerves.

HOWEVER...Chordin, Noggin, and/or Follistatin prevent BMP from stimulating ectoderm.

BY THIS INHIBITION, these three proteins stimulate nerve development above notochord. Almost just like the Holy Grail inducing substance so many researchers had tried to discover; except with these differences:

    * The "primary inducer" is a combination of proteins (Chordin, Noggin, and/or Follistat)
    * These proteins act as inhibitors of an inhibition (BMP)

If "knock out" of chordin, etc. or if antisense RNA removal of chordin, noggin etc. then skin without nerves.

Anti-sense BMP RNA prevents synthesis of BMP results in very big neural tube, or even entirely neural ectoderm.

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Yet another technique was important in the discovery of chordin, etc. " protein rescue". This means meaning compensatory substitution for proteins deliberately destroyed by UV light. Such an assay consists of injecting proteins that suspected of serving some major function, into tissues that had been damaged enough by UV not to function. The logic is that any protein that can reactivate a UV damaged process must serve some important function (& was probably what the UV light inactivated) evidence that a given protein serves the function that the UV damaged.

This "rescue" approach has been used for decades as an assay of suspected molecular functions.

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Notice that in advanced courses it often is very important to understand the methods used to make a discovery, the logical reasoning that methods are based on, and their strengths and weaknesses.

Future exam questions in this course may ask you to invent a bioassay capable of discovering what chemical produces a certain embryological effect. Your answer should include what might go wrong, and either prevent discovering any chemical with the expected function, or might result in the wrong chemical being discovered.

That is more important than memorizing the names of more than the 5 or 6 most important signal proteins and genes.

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The following review paper was written by some of the best researchers on induction:

Formation and function of Spemann's organizer
by R. Harland and J. Gerhart, Annu. Rev. Cell Dev. Biol. 13: 611-67 (1997)

And it's better written than the textbook. It is NOT assigned reading, but worth looking at.

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The following is the paper about zebra fish stripe formation by pigment cell "repulsion" by electrical depolarization of the resting potential of pigment. What an amazing subject!

Pigment Pattern Formation by Contact-Dependent Depolarization
by Masafumi Inaba, Hiroaki Yamanaka, and Shigeru Kondo
Science Vol. 335 no. 6069 p. 677, 10 February 2012

 

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