Old Problem: What Conklin missed

From Lyons et al. 2017, based on Conklin's drawings

Jon Henry and Beatrice Steinert presented the journal club session on Edwin Grant Conklin's work on the slipper snail (Crepidula). Beatrice opened the session with a historical perspective focusing on Conklin's methods for The Embryology of Crepidula (1897), which are summarized in the STAT News video below (produced by Hyacinth Empinado, published February 22nd, 2017).

Conklin worked almost exclusively from fixed embryos because he couldn't keep them alive for more than a day or so and he needed to stain his samples to visualize nuclei and cell boundaries. Once he prepared his slides, he made hundreds of camera lucida sketches from them. These sketches were not only critical for recording his observations, but they were also central to seeing itself. Conklin ultimately consolidated his camera lucida sketches into final figure drawings, which were sent off to a lithographer in Germany who copied them onto lithography stones. In large part because Conklin could only see single time points in fixed embryos, it took him a long time to piece together the developmental sequence. This process of figuring out cell divisions and fates continued all the way up to the production of the published lithograph plates (Steinert 2016).

Conklin's camera lucida sketch, final drawing, and published lithograph for figure 48. Throughout the lengthy production of this figure Conklin changed his mind about the fate of the four cells outlined in blue boxes (Steinert 2016)

Jon is from University of Illnois-Urbana-Champaign, a long-time summer researcher at the MBL (he is celebrating his 40th anniversary of coming to the MBL!), an embryo ninja, and one of the first people to greet and train Embryology students by teaching them his ninja tool making skills. Along with Dede Lyons (who will be presenting at the journal club in a few weeks), he is one of the handful of experts of the great spiralian model organism that is Crepidula (Henry and Lyons 2016).

As Beatrice discussed, Conklin spent countless hours studying Crepidula development, and what he was able to accomplish is remarkable. However, because of the limitations of his technique and the available late-19th century technology, he wasn't able to see everything and was not able to appreciate all of the developmental events. Jon presented some of these oversights in his talk:

1) Polar lobe - Conklin nearly missed the existence of the polar lobe when he published his 1897 manuscript. He ended up including a footnote about the polar lobe in the printed article. The polar lobe is a structure formed at first cleavage that eventually fuses with the D quadrant. Today we know that it is a very important structure: if removed in some spiralians, the embryos don't form a dorsal-ventral axis. Interestingly, if the polar lobe is removed in Crepidula you get twining (two D quadrants that make double axes) (Henry et al. 2017).

Left: Normal embryo. Right: Polar lobe removal caused twinning
(Henry et al. 2017)

2) Ectodermal origin of mesoderm - Conklin is credited with showing that mesoderm is derived from an endodermal lineage within the D quadrant (1897). It turns out other cells also contribute to mesoderm. For example, Dede Lyons, Jon Henry, and colleagues showed that the 3a2 and 3b2 cells undergo an epithelial-mesenchymal transition (EMT) and become mesoderm being derived from an ectodermal lineage. (See Figure 9 and Movie 8, below, in Lyons et al. 2015).

3) Trochophore larvae - Conklin believed that the first quartet micromeres 1a2, 1b2, 1c2, and 1d2 (all together called "1q2", Conklin called them the "turret cells") made the ciliated prototroch cells. However, he actually seldom observed these cells divide. Because several of his colleagues had observed them dividing in other species to form this structure, he concluded that they also must make the prototroch in Crepidula. Now we know that 1q2 cells do not give rise to the prototroch in Crepidula. Instead, they flatten out and expand on the back of the head, which helps to reposition the mouth towards the ventral surface. Then they disappear *poof* (Lyons et al. 2017).

REFERENCES
Henry and Lyons (2016) Molluscan models: Crepidula fornicata
Henry et al. (2017) Establishment and activity of the D quadrant organizer in the marine gastropod Crepidula fornicata
Lyons et al. (2015) Spiralian gastrulation: germ layer formation, morphogenesis, and fate of the blastopore in the slipper snail Crepidula fornicata
Lyons et al. (2017) Morphogenesis along the animal-vegetal axis: fates of primary quartet micromere daughters in the gastropod Crepidula fornicata
Steinert (2016) "Drawing Embryos Together: Processes of Seeing Development in Crepidula fornicata" (undergraduate thesis)