Happy to announce that our paper on orphan gene evolution in bacteria dropped in PNAS yesterday!

This is a problem I've been thinking about since the start of my PhD. Very grateful for this journey, as well as All The Friends I Made Along the Way.

Also, Eid Mubarak to those who celebrate!

This talk is happening tomorrow—as part of the Protein Structure Evolution seminar!

Click here to register if you like cool science: tinyurl.com/prose-seminar2

Organized by @lacholt.bsky.social, @caro-rocha.bsky.social, @claudiaalcar.bsky.social, @zachary-ardern.bsky.social and myself.

Cool new riboseq paper. The fact that bacterial translationmaxxers lowkey never get on the canonical protein team must mean they keep getting mogged by genetic drift or deletional bias

I can't believe this seminar's been running for a year now!

Our YouTube channel is now live—you can watch the recordings of genuinely some of the best molecular evolution talks on the internet.

tinyurl.com/ProSE-videos

(Maybe we should do a tier list soon...)

Love this new review from @lacholt.bsky.social and @bornberglab.bsky.social: www.nature.com/articles/s41...

What I really like about this group's work is they focus on the structural evolution of new genes, even though that problem is terribly challenging to address with the tech we have today

Come one, come all to the first Protein Structure Evolution talk of the year (Feb 10)!

Click here to register if you like cool science: tinyurl.com/prose-seminar2

Jointly organized by
@lacholt.bsky.social, @caro-rocha.bsky.social, @claudiaalcar.bsky.social, @zachary-ardern.bsky.social and myself.

Another month, another awesome Protein Structure Evolution talk! Register here if you like cool science: tinyurl.com/prose-seminar2

Jointly organized by @lacholt.bsky.social, @caro-rocha.bsky.social, @claudiaalcar.bsky.social, @zachary-ardern.bsky.social and myself.

(Starts in half an hour!)

In case y'all are interested in cool science about protein evolution—join our monthly seminar series! tinyurl.com/prose-seminar2

Organized by @lacholt.bsky.social @caro-rocha.bsky.social @claudiaalcar.bsky.social and myself.

(Stephen Fried is set to deliver this week's seminar in half an hour!)

@zachary-ardern.bsky.social and myself suggested just that hypothesis in our commentary on a yeast paper from the @carvunis.bsky.social lab—I don't think this has been tested yet!

Generally, I think by "non-functional" people mean "when you knock out that one protein, it doesn't affect fitness"

I understand "pervasive/spurious translation" to mean "the protein is non-functional". See the example below from Stringer et al (2021)—I think this is the common usage.

Do people disagree with this? Are there other ways of using this phrase in the literature that I'm not familiar with?

Unannotated translation products are widespread in model E. coli Genomes contain orders of magnitude more open reading frames (ORFs) than known protein coding genes, and recent work suggests there may be unannotated proteins present in even the best studied organisms. To address this gap, we used a high throughput reverse genetic toolkit to construct precise C-terminal fusions of a reporter (and control) to >120,000 ORFs in model E. coli . We found hundreds of unannotated significant hits, and individually detected >50 novel polypeptides by western blot, including ORFs within tRNA loci. Many ORFs overlap annotated genes in the sense orientation, and we found these are likely chimeric polypeptides produced by ribosomal frameshifting. Using degron based knockdowns, we identified unannotated proteins that have putative fitness effects, and we found a novel small protein that displays phenotypes consistent with a role in the mRNA degradosome. The observation of a range of unannotated translation products should lead to better annotation and understanding of the bacterial domain of life and motivates the continued exploration of genomes broadly. ### Competing Interest Statement The authors have declared no competing interest.

Unannotated translation products are widespread in model E. coli | bioRxiv www.biorxiv.org/content/10.1101/2025.09....

But I guess if the goal is to fill in gaps in annotation for less studied organisms, mass spec can be of some help. Here's a sampling of proteogenomics-based studies that makes the general point.

(Table from the same paper) (4/4)

In cases where people have tried to validate RiboSeq-supported non-annotated proteins with mass spec in the same study—generally, very few could be confirmed.

See the highlighted rows from the table below, taken from our recent review paper: genome.cshlp.org/content/earl... (3/4)

Similar conclusions have previously been reached by Fijalkowski et al (2021), and for eukaryotes, Wacholder & Carvunis @carvunis.bsky.social (2023).

These proteins are either way too small, or not expressed enough, or generate "bad" (hard to detect) peptides—mass spec doesn't like them. (2/4)

After testing virtually all reasonable permutations of sample prep, spectra acquisition, database selection, data processing...

This team (convincingly, I think) demonstrates that ~85% of the small, *annotated* bacterial proteins are undetectable by mass spec—much less non-annotated ones. (1/4) 🧵

Promoter recruitment drives the emergence of proto-genes in a long-term evolution experiment with Escherichia coli De novo gene birth from non-genic sequence has received considerable attention due to the widespread occurrence of genes that are unique to particular species. This study shows that within the short e...

We also caught such events of novel expression "in the act": i.e., in the context of experimental evolution.

Bacterial systems provide the opportunity to track fine-grained steps in gene emergence, step-by-step.

Read the paper here -

journals.plos.org/plosbiology/...

...How inert, non-expressed parts of the genomes can gain expression.

This proposed first step of gene birth has been studied extensively in bacteria, with no sign of stopping—see attached table for a sampling.

In our previous work... (6/7)

If you couldn't tell, these hypotheses are essentially unfalsifiable!

(In a manuscript currently under review, we propose a novel approach to possibly crack this dilemma)

But we also point to other, more fruitful avenues to direct new gene research in bacteria.

E.g.: (5/7)

...This question is impossible to settle!

At least, based on how the two theories and their predictions have been set up:

The "native" origin: Orphan genes evolve so fast that homology gets lost;

While "foreign" origin: They're from sources we haven't sampled yet. (4/7)

One, they're highly diverged versions of coding or non-coding sequences already "native" to the genome,

Two, they're "foreign" genes—deposited in their current habitats from sources unknown.

We review the evidence for and against these hypotheses, and conclude... (3/7)

Standard modes of thinking about gene birth—duplication via divergence—don't really apply to "orphan genes", which lack homologs.

What's more, gene birth via duplication isn't even common in bacteria.

So where did these genes come from?

Literature proposes two answers. (2/7)

Where do "orphan genes" in bacteria come from—and how do we know? (Do we even?)

I've been thinking about this problem over the course of my PhD, and our review paper is now out in Genome Research!

genome.cshlp.org/content/earl...

Here are some plot points (1/7) 🧵

I was looking for this paper but couldn't remember the exact title, so I googled "effective population size for complete dumbasses" and it worked

I got my PhD commencement at @utaustin.bsky.social before GTA 6

If you enjoy cool science about the origin of life, join us May 13 (9 AM central time). Registration link - tinyurl.com/prose-seminar2

Having followed Dr. Hlouchová's work for a long time, I can say with confidence that this will be one of the most exciting episodes we've had yet.

Cool new seminar series alert: in tomorrow's session (March 11, 3PM GMT), @zachary-ardern.bsky.social will be discussing long-term trends in protein structure evolution.

To sign up for the series: tinyurl.com/prose-seminar2

@lacholt.bsky.social @caro-rocha.bsky.social @claudiaalcar.bsky.social

Thanks so much boss!

@idanfrumkin.bsky.social (why can't I tag you?)

I don't quite understand how this happened, but I recently completed my PhD at UT Austin! On that note, I thought I should acknowledge what I think are some of the best papers in my field of research - De Novo gene birth.

@vakirlis.bsky.social @caraweisman.bsky.social @carvunis.bsky.social

This paper from the @carvunis.bsky.social lab went under my radar for a while, likely because it seems to report "negative" results. I've since realized this is essential reading for anyone attempting to detect novel proteins using mass spectrometry. Definitely cleared up a lot of my misgivings!