I asked the lead author of the Gonium genome paper, Erik Hanschen, a few questions, and he kindly agreed to let me to post his responses (see also here, here, and here). I have edited his responses only for formatting:

What led you to suspect rb involvement in multicellularity?

We started to suspect that cell cycle regulation might be involved in the evolution of multicellularity based on the results of the Volvox genome and early analysis of the Gonium genome. Both species have expansions of cyclin D genes, a critical part of the cell cycle regulation pathway, compared to their unicellular relative, Chlamydomonas. We took a closer look at the rest of the cell cycle regulation pathway and noticed structural modifications to the RB gene, which led us to the experimental transformations.

What’s the significance of the fact that RB has moved into the mating locus of Gonium and Volvox?

Sex and the evolution of multicellularity are pretty tightly entwined in the volvocine algae- many aspects of sex, anisogamy, oogamy, internal/external fertilization, anisogamy ration, all correlate with size. On the molecular side, RB has moved into the mating locus of Gonium and Volvox, and Volvox has male/female specific alleles. Now, we don’t know all the specific details- if moving into the mating locus and evolving male/female specific alleles in Volvox are a cause of increasing morphological complexity, or rather a consequence of being in non-recombining regions. The Volvox mating type locus showed there are differences in RB intron splicing both between male/female alleles and asexual/sexual expression, suggesting that RB is playing different roles in males and females and involved in sexual development. If RB is playing a role in sexual development, there’s strong selection pressure to end up in the mating type locus, which is why RB moving into the mating locus is interesting. But this isn’t yet conclusive and there’s a lot more to be understood in how multicellularity and sex relate in the volvocine algae.

Could RB have a role in anisogamy?

RB could be playing a role in anisogamy; RB alleles in Volvox are spliced differently which suggests a role in sexual development, which might include RB being responsible for the mechanistic basis for anisogamy. But these are mostly hints so far- it’s possible that RB doesn’t play a role in sexual development or anisogamy; we don’t yet conclusively know.

I thought I remembered from Volvox 2015 that the rb-transformed Chlamy had cytoplasmic bridges, is that right?

Unfortunately, we haven’t been able to look closely at the cytoplasmic bridges in RB-transformed Chlamydomonas. Tara Marriage, a postdoctoral fellow at Kansas State University, has done some fascinating research on fascilin-like genes in Gonium, but I’d prefer not to steal her thunder.

If so, do you have an idea of how rb causes cytoplasmic bridges?

It looks like cell cycle regulation, through RB, has been co-opted to cause development of colonies- RB might be regulating cell-cell adhesion genes in a different way in Gonium than Chlamydomonas does, thus causing cells to stick together.

It’s not clear to me how exactly rb is causing cells to stick together; can you explain that?

There’s definitely more work to be done here, but one pathway would be the RB gene in Gonium is regulating downstream genes, which would likely include cell-cell adhesion genes, in a different way than the RB gene in Chlamydomonas does.