What I’ve learned about water, aging and protein quality control

When I was a biology student at the Chinese University of Hong Kong, I learned about living organisms, cells and their building blocks such as nucleic acid and proteins. I do not recall a focused discussion about the water molecule or the importance of water to life.

Alice Liu

Water is simple, ubiquitous and taken for granted. I did not appreciate the importance of water in biology and life until late in my science career.

I have always been interested in quality control, or QC. In biology, molecular chaperones are important for the proper folding and quality control of our proteome, particularly under stress. Not surprisingly, transient hyperthermia, or overheating, increases the expression of these protein chaperones, thus the term “heat shock proteins,” or HSPs. In the early days of my work, I reasoned that the dysfunction of this protein QC mechanism likely contributed to age-related dys-homeostasis and vulnerability to diseases. 

My lab’s first publication on this appeared in the Journal of Biological Chemistry in 1989. This and related studies from other labs on cells, animals, and human peripheral lymphocytes suggest that the proteome declines with age and under stress.  We hypothesized HSPs helped maintain protein homeostasis and diminished HSP production would contribute to age-related deterioration and vulnerability to diseases, in particular, neurodegenerative diseases, such as Huntington’s that are known for  disease protein aggregates.   

When we acquired a cell model that expressed a reporter disease protein of Huntington’s disease, or HD, we tested whether a heat shock–induced increase in HSP chaperones — which support protein structure and function in cells under stress — would prevent the misfolding and aggregation of the disease protein into what are called inclusion bodies, or IBs.

To our surprise, HS and HSP chaperone expression increased the abundance of IBs at the expense of the soluble and diffusible form of the reporter protein. We could not make sense of all these round, fluorescent-green, micron-size entities in our heat-shocked cells. The result was the opposite of what we expected.  

We combed the literature to figure out what it meant, and we quickly realized that a protein's amino acid sequence determines its folded 3D structure, which then determines its biological function.

Evidence has been accumulating for more than two decades that 30% to 50% of the proteome of higher eukaryotes, in their native and functional states, is disordered and unfolded, entirely or in parts. Their amino acid composition — low in hydrophobic and high in charged amino acid residues — lacks the driving force of hydrophobic collapse needed to fold and compact into a defined 3D structure in a water-based cell environment.

These natively unfolded, functional proteins are termed intrinsically disordered proteins and proteins with disordered regions. They are exquisitely sensitive to cell crowding and changes in cell hydration and are overrepresented among regulatory proteins including proteins implicated in cancer and neurodegenerative disease, prompting what researchers called the “disorder in disorders” or “D2” concept to underscore this intimate association of structural disorder of disease proteins and disease pathogenesis.

Alice Liu

Students in Alice Liu’s lab include, in back, Praneet Paruchuri and Sachin Mysore and, in front, Myera Mlan, Kaylie Lam, Amala Mathew and Rida Shah. Mysore and Paruchuri are co-authors of the lab’s 2023 ACS Chemical Neuroscience paper, and Mathew is a co-author a recently published book chapter.

That reporter disease protein of Huntington’s we’d studied is intrinsically disordered in structure.

Many neurodegenerative diseases are known to be age related. Symptoms of autosomal dominant diseases such as HD show up late in life, reflecting the age-related changes in the cell environment that allow the disease to progress. Both lay and scientific evidence shows that aging — from yeast to human — is associated with decreased hydration and changes in water homeostasis.

The human brain’s structure and function make it particularly vulnerable to such age-related changes. Our rigid skull limits how much tissue can expand or shrink, and neurons lack aquaporin for water transport. Further, many compatible organic osmolytes for tonicity and volume control in peripheral tissues such as the kidney medulla and marine organisms, including glycine, GABA, taurine and glutamate, as well as potassium, sodium and chloride, are essential for neurotransmission, precluding their use as osmo-regulators to counter shrinkage and restore volume in neurons.   

In short: Water, water everywhere, but not a drop for the sickly neurons of the aging brain.

In our study of how the mutant HD proteins form aggregates in cells, we saw the pivotal role of water in cell function versus dysfunction and became aware of the role age-related changes in hydration can play in regulating the structure and function of our proteome — from the stably folded to the intrinsically disordered.

The delicately balanced give-and-take between hydration and structural dynamics governs a protein’s function versus dysfunction in a cell; the strength of the coupling with water across different protein classes decreases from the structurally dynamic and disordered to the folded proteins.

A hypo-hydrated and crowded aging cell environment would promote the structuring, compaction and aggregation we saw for the mutant HD reporter protein. And this environment also would bolster the compact and folded structure of the folded proteome to dampen the need for help from HSP chaperones under stress that we reported more than 30 years ago.  

As Yaakov Levy and José Onuchic wrote in a 2004 commentary, “fluctuations of the hydration water can slave the protein dynamics and thus affect its function.” 

I belatedly became interested in water because I needed to understand and resolve the conundrum of a time-delayed and age-dependent manifestation of an autosomal dominant neurodegenerative disease trait. This reminds me that studies of biological processes must be grounded in the context of our water-based and crowded cell environment. It has been an unexpected and rewarding journey.  

I am thankful that I have a chemist as my research partner — Kuang Yu Chen, professor emeritus of chemistry and chemical biology at Rutgers University — who I can turn to for chemistry-related issues. Without his patience and help, I doubt I would have gone this far in this work. And this body of work would not have been possible without the contributions of my colleagues, postdocs and students over the years. I am indebted.  

These days, in my classroom teaching of cell biology, I strive to emphasize the importance of learning fundamental concepts well — most importantly of water, the molecule for life as we know it — to better understand and appreciate life processes.