Bonkers Really … but …

 

This is just in case you spotted the headline in January 2018: ‘Scientists Counted All The Protein Molecules in a Cell And The Answer Really Is 42. This is so perfect.’ 

Them scientists eh! The things they get up to!! The scallywags in this case were Brandon Ho & chums from the University of Toronto and Signe Dean, the journalist who came up with the headline, was referring, of course, to Douglas Adams’s “Answer to the Ultimate Question of Life …” in The Hitchhiker’s Guide to the Galaxy — though it may be noted that Ho’s paper includes neither the number 42 nor mention of Douglas Adams.

The cult that has evolved around this number is both amusing and bizarre, not least because Adams himself explained that he dreamed 42 up out of the blue. In a different context a while ago (talking about how the way you get to work might affect your life expectancy) I recounted happy evenings spent carousing in The Baron (well, having a quiet jar or two) with Douglas Adams and friends from which it was clear that he was not into abstruse mathematics, astrology or the occult. He just had a vivid imagination.

Anything for a catchy headline but

Aside from the whimsy, is there anything interesting in this paper? Well, yes. Ho & Co studied a type of yeast (Saccharomyces cerevisiae) that is mighty important because it’s been a foundation for brewing and baking since ancient times. So no merry sessions in The Baron of Beef without it! Its cells are about the same size as red blood cells (5–10 microns in diameter) but you can actually see them sometimes as films on the skin of fruit. It’s played a huge role in biology as a ‘model organism’ for studying how we work because the proteins it makes that are essential for life are pretty well identical to those in human cells — so much so that you can swap those that control cell growth and division between the two. Yeast proteins work just fine in human cells and vice versa.

 

Yeast on the skin of a grape. Photo: Barbara W. Beacham

 

The question Ho & Co asked was ‘how many protein molecules are there in one cell?’ In the age when you can sequence the DNA of practically anything at the drop of a hat, you might think we’d know the answer already but in fact it’s not been at all clear. Accordingly, what these authors did was to pull together all the relevant studies that have been done to come up with an absolute figure. The answer that emerged was that the number of protein molecules per yeast cell is 4.2 x 107 — which, of course, can also be written as 42 million. Eureka! We have our headline!! Albeit, as the authors noted, with a two-fold error range.

Does anyone care?

Now you’re just being awkward. You should be grateful to be made to picture for a moment tens of millions of proteins jiggling around in little sacs so small you could get tens of thousands of these cells on the head of a pin. And somehow, in that heaving molecular city, each protein manages to carry out its own task so that the cell works. It is quite staggering.

Mention of tasks leads to the other question Ho et al looked at: how many copies are there of the different types of protein? We know from its DNA sequence that this yeast has about 6,000 genes (Saccharomyces Genome Database). So that’s at least 6,000 different proteins. Not surprisingly, it turns out that about two thirds of them are in the middle in terms of abundance — i.e. there’s between 1,000 and 10,000 molecules of each sort per cell. The rest are either low abundance (up to about 800 molecules per cell) or at the high end — 140,000 to 750,000, i.e. somewhere in the region of half a million copies of each type of protein.

Does this distribution make sense in terms of what these proteins do?

You know the answer because if it didn’t the Toronto team wouldn’t have got their work published but, indeed, proteins present in large numbers are, for example, part of the machinery that makes new proteins (so they’re slaving away all the time) whereas, those present in small numbers do things like repair and replicate DNA and drive cells to divide — important jobs but ones that are only intermittently needed.

These results aren’t going to turn science on its head but it is awe-inspiring when a piece of work really brings us face-to-face with stunning complexity of biology. And if it takes a bonkers headline to catch our eye, so be it!

Reference

Ho, B. et al. (2018). Unification of Protein Abundance Datasets Yields a Quantitative Saccharomyces cerevisiae Proteome. Cell Systems. Published online: January 23, 2018.

Hares And Tortoises

You may have noticed that the last few months have seen a bit of a DNA-fest in these pages. Don’t blame me. It’s all the fault of them scientists beavering away in their labs. We’ve just done “Making Movies in DNA“, in “And Now There Are Six!!” the genetic code was expanded from four to six units by making two new ones artificially and in “How Does DNA Do It?” we saw how words can be transformed into a sequence of DNA.

Now they’re at it again – or at least Stephen Kowalczykowski, James Graham and colleagues of the University of California at Davis are – revealing yet more astonishing things about this molecule, just when you could be thinking we’ve got the hang of it.

I might add that I’m grateful to my correspondent David Archer of The Society of Biology for bringing this piece of work to my notice as I’d missed it in the journal Cell (cries of ‘shame’ and ‘shurely shome mistake’ mingle in the background).

What is it this time?  

Well it’s two really astonishing things about DNA replication – the process by which double-stranded DNA is pulled apart so that each strand can act as a template for making a new DNA molecule. Result: as cells progress towards division, they double their DNA content so that equal amounts can be given to each new daughter cell. The first source of amazement is that Stephen K & chums have filmed this happening in real time. That’s a terrific feat – but what it reveals is quite bizarre.

Up to now it’s been assumed that the protein machines (DNA polymerases) doing the biz trundle along each of the separated strands of parental DNA at more or less the same speed. It would seem to make no sense to do otherwise and risk ending up with the job half done. In other words, the duplication of the two strands is coordinated. Is that what K & Co found? Not a bit of it! Extraordinary to relate, it appears that there’s no coordination between the strands at all!! Not for the first time in the history of molecular biology a technical advance has thrown up the totally unexpected. Before we look at the results in a bit more detail, a little background might be useful.

One divides into two

Making two identical copies of DNA from one original happens every time one cell divides to make two. And there’s a lot of it about. As is well known, we all start out as one cell (i.e. a fertilized egg) that turns into a human being – 50 trillion cells (that’s 5 + 13 zeroes). And even after we’ve been assembled it takes a lot of cell-making to keep us ticking over – about one million new cells every second. Just take a second to think about that: DNA comes in the well-known form of a double helix – two strands made up of chemical units (called nucleotides) linked together. Each unit has one of four bases (cytosine (C), guanine (G), adenine (A), or thymine (T)) and the strands are “complementary” because C pairs with G and A with T – a rigid rule that means if you know the sequence of bases in one strand you can work out what it is in the other. So far so simple. But, as we noted in “How Does DNA Do It?”, the coding power of DNA lies in its size. In us three billion letters are available to do the encoding. That is, there are just over 3,000 million units in each chain – i.e. 3,000 million base-pairs all told. And all of these are copied (twice) for every new cell.

DNA replication: The double helix is ‘unzipped’ so that each separated strand (turquoise) can act as a template for replicating a new partner strand (green). This creates a ‘replication fork’ – two branches of single stranded DNA. The new strands are made by protein complexes called DNA polymerases chugging along the parent strands, making new, complementary, strands as they go. There’s a small technical wrinkle here: new DNA chains can only be extended in one direction. This means that, while one strand can be made continuously (the leading strand), the other has to be put together in short bits as the parent strand is unwound, with the bits being joined up afterwards (the lagging strand).

 

 

Timing is everything

So the cell’s task is to unzip the double helix and use each exposed strand as a template for building a new partner strand. Things are helped by DNA being split into fragments (chromosomes: 23 pairs in humans + 2 sex chromosomes, 46 per cell all told). Even so, chromosomes are huge: the longest (chromosome 1) has nearly 250 million base-pairs; the shortest (chr 21) has about 47 million. The problem for the machinery that has evolved for the job is that it cranks along at 50 pairs per second – roughly a month per chromosome. But in a normal cell cycle the whole business is done in about two hours! That’s made possible because replication doesn’t do the obvious: start at one end and work its way to the other. Cunningly it hits lots of ‘start points’ – up to 100,000 in a single cell – making lots of short bits at the same time that are then joined up. In other words replication proceeds simultaneously from many different sites in chromosomes. Enzymes join the pieces together to make the final, complete copy.

It’s rather like you having some horribly repetitive chore to do – washing up after a big dinner. On your own you might start at one end of the pile and work through it but, far better, get one member of the family to do the plates, another the cutlery, etc. and – job done!!

Now for today’s bit of amazing science

What Kowalczykowski and friends did was to extract DNA from bugs (E.coli bacteria in fact, that can make DNA about 20 times faster than human cells), set up a replication system and measure what went on by microscopy, using a dye (SYTOX Orange, which is fluorescent) that sticks to complete double helices but not to single strands. Thus they could track progress along a strand as a new double helix formed. What they saw was that each strand acted independently of the other. Overall, the rate of replication of the two strands was about the same (as it must be in the end) but along the way there were stops and starts and sometimes one strand would grow at ten times the speed of the other. How weird is that?!!

Seeing DNA being made. In this picture microscopy reveals three extending stretches of double-stranded DNA being made (Graham et al. 2017). Click here to see video.

You could picture DNA replication as one of those Swiss railway trains cranking up a mountain at an improbable angle, using a rack-and-pinion to stop it sliding backwards. Think of the engaging cogs as new base-pairs. The train just keeps chugging along until it reaches the its next stop. But why doesn’t the DNA-making machinery do the same? Well, we haven’t much of a clue. One difference is that the train has its track (and rack) laid out before it, whereas DNA is continuously being unwound to open the template. Some bits are more difficult to unwind than others and this variation may cause the system to go in fits and starts. Another contribution many come from the many proteins involved in this complicated process. As well as the polymerases there are things that unwind DNA, stabilize it, stitch new bits together, etc. and these complexes are continuously forming, falling apart and re-assembling – all of which gives plenty of scope for erratic behaviour.

Fact of the matter is, we don’t know. So, in revealing completely unexpected behaviour, this technical triumph throws up the question of how two strands working independently manage, in the end, to come up with the perfect finished product.

But hey! This wouldn’t be science if we had all the answers!

Reference

Graham, J.E., Marians, K.J., Kowalczykowski, S.C. (2017). Independent and Stochastic Action of DNA Polymerases in the Replisome. Cell 169, 1201–1213.