DEVO-7: Gradients & Morphogens –1–

Review and move on… DEVO-7

• Complete Left-right asymmetry paper • On to morphogensis (gradients, responses, and patterns)

Q: A question (from you) about the myoD1 paper? Types of –2–

Q: what types of are you familiar with?

Q: How might these different myosins have arisen (evolutionarily)?

DEVO-7 Question from the past –3–

Q: why does muscle contract Q: why is it that MTs can support rather than expand (or both)? movement in both directions?

DEVO-7 Cellular handedness –4–

Q: how do and systems interact?

DEVO-7 Cellular handedness –5–

Q: Does the microtubule system have a handedness?

DEVO-7 Thinking about myosin 1D / 1C paper –6–

we will be reasonably superficial

• What did they know before they started? • What was the author’s assay system - what were they looking at? • What did they do? • What did they find?

DEVO-7 Types of myosins –7–

myo1D is (one of a number of) situs inversus : its absence leads to the full reversal of organ positioning along the LR axis, with organs adopting a mirror-image orientation.

DEVO-7 (A) Control larvae showing bilateral symmetry. The trachea are indicated, with anterior tracheal spiracles indicated by a red (left) or blue (right) (Bottom) Front view of the larvae (ventral down, dorsal up).

MH, mouth hooks; ALS, anterior left spiracle; ARS, anterior right spiracle; PLS, posterior left spiracle; PRS, posterior right spiracle.

A – anterior; P – posterior; L – left; R – right; D – dorsal; V– ventral. “we ectopically expressed the in different naïve tissues (i.e., tissues devoid of LR asymmetry). We found that Myo1D expression in the larval epidermis induces dextral twisting of the whole larval body

Q: what is ectopic expression and how did they do it. Q: what does it mean that “phenotype is 100% penetrant and is specific to Myo1D”?

DEVO-7 Linear morphology of a control (btl>srcGFP) third myo1D expression in the instar larvae trachea. tracheae (btl>myo1D) increases their dextral twisting with Q: What does GFP multiple loops. expression control for? Co-expression of myo1D and an RNAi against myo1D completely suppresses the twisting phenotype

Q: what does this experiment “control” for? is a positive or negative control?

DEVO-7 myo1D mRNA levels (by qPCR) in the epidermis of control (tsh>), myo1D-GOF (gain of function) (tsh>Myo1D) or myo1C-GOF larvae (tsh>Myo1C). Error bars indicate standard deviation, RNA levels in arbitrary units (AU).

Q: how did they get these data and why did they include it? Myo1C is a sinistral myosin, an antagonist to Myo1D.

Control larvae (A) and larval-twisting phenotype (B) induced by over- expression of myo1D or by the over-expression of (C) myo1C or the co- expression of (D) myo1D & myo1C.

DEVO-7 Q: What is a chimera? Myo1D:Myo1C chimeras indicate that head/neck domain provides directionality to the . Domains designated D & C derived from Myo1D and Myo1C, respectively.

chimeras

DEVO-7 myosin handedness –17–

Q: How would the presence or absence of ATP influence this experiment?

DEVO-7 • At the adherens junction (↑) MyoID–β- interaction is stabilized cadherin. MyoID triggers dextral development. • B: MyoIC over-expression (→) disrupts MyoID-β-catenin interaction and MyoID’s function leading to sinistral development. • D: In the absence of DE-cadherin (→), there is no MyoID over-expression rotation phenotype. • What is missing from the “model”? Q: What does a morpholino do?

Organ situs of wild-type (A) & myo1d morphant tadpoles displaying situs inversus (B) and heterotaxia (C) at stage 45. D: Quantification of organ situs analysis. Numbers of analyzed specimens, derived from 3 (D) independent experiments. Your question –20–

Q: Major questions (from you) about the morphogen paper?

DEVO-7 FIRST

Q: what is going on here? what do the arrows and bars mean?

Q: what would you need to know to predict the behavior of such a DEVO-7 system? Morphogens –24–

https://www.ibiology.org/development-and-stem-cells/enhancers/#part-1

background through 7:29 Next : Complete morphogens and: Genikhovish & Technau. 2017 On the evolution of bilaterality