living cell (4)

Vesicles shuttle proteins and membranes between intracellular compartments, allowing cells to eat, drink and secrete. [p. 511]  Taking a closer look [below figure 15-22] we see that the docking process is quite involved.  Also note that it is critical that the vesicle dock on the appropriate target membrane.  

To function optimally, each transport vesicle that buds off from a compartment must take with it only the proteins appropriate to its destination and must fuse only with the appropriate target membrane.  A vesicle carrying cargo from the Golgi apparatus to the plasma membrane, for example, must exclude proteins that are to stay in the Golgi apparatus, and it must fuse only with the plasma membrane and not with any other organelle.  [p. 512]

This means that the cargo is very specific, and that somehow this specific cargo can be selected, sorted, bundled and shipped to a precise location within the cell.  Recall our altered perspective: if the vesicle is five or six inches in diameter, the diameter of the cell is something like 4000 feet and over 260 billion cubic feet.  Navigation from one point to another in this crowded environment is not simple.

But we don’t believe that the vesicles simply float in the cytosol from one place to another.  In fact, they get carried by a special protein called kinesins.  Kinesins are a form of motor protein capable of moving [see below in figure 4-50].

These kinesins (and dyneins) carry vesicles along microtubules.  Microtubules serve several functions within a cell, but in this instance provide an internal highway system for intra-cellular transport.  Microtubules are themselves proteins and are being constantly assembled and disassembled throughout the interior of the cell.  

As can be seen below [figure 17-20] these various kinesins and dyneins ‘walk’ along a microtubule to deliver their vesicle to the proper location within the cell.

So how does the originating location (the Golgi apparatus, for instance) ‘know’ which microtubule leads to the desired location?  After all, the cell is full of microtubules going several different places.  For that matter, how does the Golgi apparatus ‘know’ which location to send a particular bundle of proteins in the first place?  Or to take it back one more step, how does the Golgi apparatus ‘know’ that a particular location within the cell requires a particular set of proteins?

Consider the kinesins themselves.  If we saw one the size of a grasshopper striding along a garden path we would immediately recognize it as something utterly different than the rocks in the same garden.  We would think, ‘I don’t know that thing is, but I know that it is alive.’  In the same way, if we saw a ribosome the size of a grapefruit spitting out a lengthy string of amino acids the size of raisons we would think, ‘What an interesting little machine!’  When that string of raisons began, with the help of other strings of raisons, to form into a particular shape, and then merged spontaneously with other stringed raisons, we would know we are witnessing something different from the rain or the running of a river.

It lies beyond human credulity to think that the biological processes necessary to synthesize proteins came about by chance (as Dawkins suggests) or that it operates entirely utilizing the physical and chemical forces and properties we currently understand.  These are living processes, and differ from the non-living chemical processes in the same way a swordfish differs from a stone.  One is alive, and the other isn’t.

DNA replication requires the cooperation of a large number of proteins that act in concert to synthesize new DNA.  These proteins form part of a remarkably complex replication machine. [p. 211]

Protein synthesis is only one of many remarkable biological processes that exist within a living cell.  Even so, there is probably none that are as central to life itself, or amazing in its complexity and mystery, than cell division and DNA replication.  All life is cellular; the only way to make a cell is via another cell, and the only way another cell can thrive is through replicated DNA.  

One of the interesting facts about DNA is how much of it seems to be junk and unusable, while other non-coding segments seem crucial yet without known functions, as there is 

…a large excess of interspersed DNA.  This extra DNA is sometimes erroneously called ‘junk DNA’ because its usefulness to the cell has not yet been demonstrated.  Although spare DNA does not code for a protein, much of it may serve some other biological function.  Comparisons of the genome sequences from many different species reveal that small portions of this extra DNA are highly conserved among related species, suggesting their importance for these organisms. [p. 180]

Before examining DNA replication in detail, a quick overview of cell division will provide the necessary context.  The figure below [18-2] lays out the four major stages.  DNA replication takes place during S phase.  During the gap phases (G1 and G2)...

...the cell monitors both its internal state and external environment.  This monitoring ensures that conditions are suitable for reproduction and that preparations are complete before the cell commits to the major upheavals of S phase…and mitosis.  At particular points in G1 and G2, the cell decides whether to proceed to the next phase or pause to allow more time to prepare. [p. 611]

So somehow, something within the cell ‘decides’ whether to proceed to the next phase.  What decides?  And how does the deciding entity ‘know’ the conditions throughout the entire cell?

This understanding (the conditions within the cell) is crucial, because

At the transition from G1 to S phase, the control system confirms that the environment is favorable for proliferation before committing to DNA replication.  Cell proliferation in animals requires both sufficient nutrients and specific signal molecules in the extracellular environment; if these extracellular conditions are unfavorable, cells can delay progress through G1 and may even enter a specialized resting state.

At the transition from G2 to M phase, the control system confirms that the DNA is undamaged and fully replicated, ensuring that the cell does not enter mitosis unless its DNA is intact.

This suggests that the cell has a way to interpret external and internal signals to ensure that the time is right for proliferation.  In particular, the state of DNA replication.  As we examine DNA replication closer, it will become apparent how difficult it must be to determine the precise condition of all the DNA and the integrity of its duplication.  But this is only one part of the challenge:

Two types of machinery are involved in cell division: one manufactures the new components of the growing cell, and another hauls the components into their correct places and partitions them appropriately.  When the cell divides in two, the cell-cycle control system switches all this machinery on and off at the correct times, thereby coordinating the various steps of the cycle.  The core of the cell-cycle control system is a series of molecular switches that operate in a defined sequence and orchestrate the main events of the cycle, including DNA replication and the segregation of duplicated chromosomes. [p. 613]

‘Types of machinery’, ‘hauls components’, ‘correct places’, ‘control system’, ‘molecular switches’, ‘machinery’, ‘correct times’, ‘coordinating’, ‘defined sequence’, and ‘orchestrate’.  All this life-like, intentional, and organized activity indicates that we are not talking about advanced chemistry.  But it becomes even more amazing when we delve into DNA replication.

Nothing is more central to life than DNA.  DNA is the source of all the intelligence, information, control, and architecture that we are aware of.  And not just within a living cell: within the entire organism, and in some respects, entire societies, if we believe that human nature arises from human DNA.  And nothing is more central to the continuation of life than the replication of DNA.

Because each strand of DNA is complementary, it acts as a template for its replication. (see below figure 6-2)

As part of cell division, the DNA is replicated in S phase at a rate of about 1000 nucleotides per second [p. 200].  The process of DNA replication is similar to RNA transcription we reviewed earlier, with additional complicating factors.  For example:

A serious problem arises…as the replication fork approaches the end of a chromosome:  although the leading strand can be replicated all the way to the chromosome tip, the lagging strand cannot.  When the final RNA primer on the lagging strand is removed, there is no enzyme that can replace it with DNA.  Without a strategy to deal with this problem, the lagging strand would become shorter with each round of DNA replication, and, after repeated cell divisions, the chromosomes themselves would shrink – eventually losing valuable genetic information. [p. 213]

A DNA polymerase is the roving factory that performs the replication of DNA, with special processes for each end [see figure 6-20 below].  

As can be seen in the figure above, DNA replication is a complex and coordinated process, one that includes several different proteins, as can be seen in the table below [6-1]:

Instead of tracking the process in detail, I will focus on a few interesting complexities, in particular error correction and DNA bundling:

Although simple in principle, the process [replicating DNA] is awe-inspiring, as it can involve the copying of billions of nucleotides pairs with incredible speed and accuracy: a human cell undergoing division will copy the equivalent of 1000 books like this one [Essential Cell Biology] in about 8 hours and, on average, get no more than a few letters wrong. [p. 200]

According to the text, “Polymerization and proofreading are tightly coordinated, and the two reactions are carried out by different domains in the same polymerase molecule.” [p. 208].  

We see in this molecular machine (DNA polymerase) performing several coordinated functions in order to ensure the DNA is right:

DNA polymerase is so accurate that it makes only about one error in every 104 nucleotide pairs it copies.  This error rate is much lower than can be explained simply by the accuracy of complementary base pairing…in correct base pairs are formed much less frequently than correct ones, but, if allowed to remain, they would result in an accumulation of mutations.  This disaster is avoided because DNA polymerase has two special qualities that greatly increase the accuracy of DNA replication.  First, the enzyme carefully monitors the base-pairing between each incoming nucleoside triphosphate and the template strand.  Only when they match is correct does DNA polymerase undergo a small structural rearrangement that allows it to catalyze the nucleotide-addition reaction.  Second, when DNA polymerase does make a rare mistake and adds the wrong nucleotide, it can correct the error through an activity called proof reading. [p. 208]

There are two processes to limit mistakes in DNA replication.  As mentioned above, the first is proof reading:

Proof reading takes place at the same time as DNA synthesis.  Before the enzyme adds the next nucleotide to a growing DNA strand, it checks whether the previously added nucleotide is correctly base-paired to the template strand.  If so, the polymerase adds the next nucleotide; if not, the polymerase clips off the mis-paired nucleotide and tries again. [p. 208]

Again we see a deliberate, intentional and meaningful process conducted by complex macromolecules.  Can we continue to attribute this solely to chemistry and purposeless macromolecules?

While errors are rare with proofreading, when they occur the backup process known as mismatch repair takes place. [see figure 6-29 below]

As a reminder, DNA replication takes place in phase S of cell division.  Once DNA replication is complete, the DNA must be bundled.  Recall that at times other than cell division, the DNA floats around the nucleus, essentially invisible.  Also recall that if each nucleotide was the size of a golf ball, a single DNA molecule would stretch over 6000 miles. Or as the text puts it:

Wrapping all the DNA in a human cell is like taking a fine thread 24 miles long and folding it into a tennis ball. [p. 178]

Understanding the scale of DNA bundling is important to grasp how incredible it is:

The complex task of packaging DNA is accomplished by specialized proteins that bind to and fold the DNA, generating a series of coils and loops that provide increasingly higher levels of organization and prevent the DNA from becoming a tangled, unmanageable mess.  Amazingly, this DNA is folded in a way that allows it to remain accessible to all of the enzymes and other proteins that replicate and repair it, and that cause the expression of its genes. [p. 178]

So in addition to the complexity of bundling the fine thread of a DNA molecule that is 24 miles long into the size of a tennis ball, it needs to keep in mind what parts of the molecule may need to be accessed:

The localized alternation of chromatin [DNA] packing by remodeling complexes and histone modification has important effects on the large-scale structure of interphase chromosomes.  Interphase [during cell division] chromatin is not uniformly packed.  instead, regions of the chromosome containing genes that are being actively expressed are generally more extended, whereas those that contain client genes are more condensed.  Thus, the detailed structure of an interphase chromosome can differ from one cell type to the next, helping determine which genes are switched on and which are shut down.  Most cell types express only about half of the genes they contain, and many of these are active only at very low levels.  

To understand this process better, and appreciate the implications of the expressed intelligence and coordination, consider the figure below [5-24]

Keep in mind the relative length of this molecule, and the challenge to make available a specific small part of it to be accessed for gene expression:

Both ATP-dependent chromatin [DNA] remodeling complexes and histone-modifying enzymes are tightly regulated.  These enzymes are often brought to particular chromatin regions by interaction with proteins that bind to a specific nucleotide sequence in the DNA – or in an RNA transcribed from this DNA.  Histone-modifying enzymes work in concert with the chromatin-remodeling complexes to condense and relax stretches of chromatin [DNA], allowing local chromatin structure to change rapidly according to the needs of the cell.  [p. 189]

How does the cell know what is needed?  How does the necessary enzymes navigate to the required strand of DNA?  Keeping in mind that many DNA molecules exist in the same nucleus, and every one of them is very very long.  

The need for this regulation is clear: in order to provide only what the cell needs, and not more, it is necessary to limit waste and to produce only what is required at a given time.  But how is this planning/coordination/navigation accomplished?